Specification

The present invention relates to compounds according to general formula (I) with enhanced absorption of UV-B irradiation. The present invention also relates to a UV-B tolerant transgenic plant and a method for enhanced production of compounds according to general formula (I) in a plant or plant cell using a transgenic plant. Furthermore, the invention relates to a nucleic acid sequence (SEQ-ID No.1 ) encoding FPT2, which catalyzes the production of compounds according to general formula (I).

The present invention therefore comprises a claim for a given product, namely the compounds according to general formula (I), an independent claim for a process especially adapted for the manufacture of these compounds, independent claims referring to a use of these compounds and in addition an independent claim referring to nucleic acid constructs and transgenic plants specifically designed for carrying out said process for manufacture of the inventive compounds. The invention further relates to compositions comprising compounds according to general formula (I). Furthermore, the invention relates to a method of conferring UV-B tolerance to a plant as well as an UV-B tolerant plant comprising the nucleic acid sequence SEQ-ID No.1 encoding FPT2, which catalyzes the production of compounds according to general formula (I).

Background of the invention

It is intended to contribute to the solution of environmental problems caused by destroying the ozone layer and serious problems concerning the food resources in the 21 ^st century. The zonal average ultraviolet irradiance reaching the Earth's surface has continuously increased since 1979 at all latitudes except the equatorial zone even though the total area of the ozone hole has slightly decreased since 2006, most likely due to efforts restricting usage of chlorofluorocarbons.

The dependency of plants upon sunlight inevitably brings them into exposure to ultraviolet light, including that in the wavelength range 280-320 nm (UV-B). This wavelength range is potentially damaging to DNA, RNA and proteins and furthermore leads to increased production of free radicals that can activate transposons and cause mutations. Thus although only 0.5% of energy reaching the earth is in this wavelength range the risk of UV- B damage is profound. The potential impact of UV-B irradiation on natural plant populations could greatly impair crop yields since the detrimental effects described above combine to considerably constrain plant seed yields and seed germination rates. For this reason, plants have been under considerable natural selective pressure to generate elegant mechanisms to both sense and to respond to the presence UV-B irradiation.

One aspect of the response in land plants is the production of additional antioxidant sunscreen pigments including flavonoids and hydroxycinnamates, which belong to phenylpropanoids.

Flavonoids constitute a relatively diverse family of aromatic molecules that are derived from phenylalanine and malonyl-coenzyme A (malonyl-CoA). In the phenylpropanoid pathway, chalcone synthase (CHS) uses malonyl-CoA and 4-coumaroyl-CoA as substrates. Afterwards, chalcone isomerase (CHI) catalyses ring closure to form a flavanone. Further enzymes in the pathway are: flavanone 3-hydroxylase (F3H), dihydroflavonol 4- reductase (DFR), flavonoid 3'-hydroxylase (F3'H) and flavonoid 3',5' hydroxylase (F3'5H). Natural variance screening has been carried out in many plant species including Arabidopsis. However, with regard to the accumulation of specific primary and secondary metabolites relatively few studies have focused on phenylpropanoid metabolism. However, plants with enhanced UV-B irradiation protection mechanisms are urgently needed, because the vast majority of people on earth are dependent on plant-based diet. Furthermore, phenolic compounds with the ability of defence against UV-B irradiation are needed as well.

Therefore, a primary object of the present invention was the provision of compounds that absorb UV-B irradiation and thereby protect biological materials as well as non-biological materials from damaging exposure to UV-B irradiation. Further object of the invention was the provision of genetically modified organisms, capable of providing compounds that absorb UV-B irradiation and thereby protect biological material from damaging exposure to UV-B irradiation. An additional object of the present invention was the provision of formulations and compositions comprising such compounds.

The objective of the present invention is solved by the teaching of the independent claims. Further advantageous features, aspects and details of the invention are evident from the dependent claims, the description, the figures, and the examples of the present application.

Description of the invention

Assessment of levels of various phenylpropanoids in flowers in a set of 72 Arabidopsis thaliana ecotypes was performed by the inventors. Application of liquid chromatography- mass spectrometry resulted in the detection of unknown chemical structures. MS-MS fragmentation studies suggested that these unknown peaks are novel flavonol derivatives. In order to characterize the chemical structure of these peaks large amounts of the C24 accession that accumulated these compounds were re-grown and purified. Column chromatography and HPLC analysis was carried out (see Example 1 and 2).

Surprisingly, compounds according to general formula (I), novel phenylacylated flavonol- glycosides, named saiginols, which display enhanced UV-B absorbent properties (Fig. 3 a- c), were detected. However, compounds according to general formula (I) do not absorb in the photosynthetically active radiation range. Accordingly, there is no trade off against photosynthetic efficiency. Furthermore, the position of the phenylacyl moieties of the compounds according to general formula (I) is structural distinct from all phenylacylated flavonols reported to date. Characterization of a compound according to general formula (I) was performed by NMR. Fig. 3d shows a computational estimation of the most stable stereochemical structure of a compound according to general formula (I).

According to an aspect of some embodiments of the present invention a compound of general formula (I) is provided

wherein

R ¹ , R ² and R ³ are independently of each other selected from: -H, -OH, and

-OCH ₃ ;

R ⁴ is selected from -H, -OH, and -OCH ₃ ;

and enantiomers, mixtures of enantiomers, diastereoisomers, mixtures of diastereoisomers, hydrates and solvates thereof. According to an aspect of some embodiments of the present invention a compound of general formula (II) is provided

wherein

R ⁴ is selected from -H, -OH, and -OCH ₃ ;

R ⁵ represents -H, or

and enantiomers, mixtures of enantiomers, diastereoisomers, mixtures of diastereoisomers, hydrates and solvates thereof.

According to an aspect of some embodiments of the present invention a compound of general formula (III) is provided

wherein

R ⁴ is selected from -H, -OH, and -OCH ₃ ;

and enantiomers, mixtures of enantiomers, diastereoisomers, mixtures of diastereoisomers, hydrates and solvates thereof.

The term "saiginols", as used herein, refers to the compounds according to the general formula (I), as given above. Since the general formulas (II) (III) and (IV) fall under general formula (I), the term "saiginols", as used herein, refers also to the compounds according to general formula (II), (III) and (IV) as given below.

According to an aspect of some embodiments of the present invention a compound of general formula (IV) is provided

wherein

R ⁴ is selected from -H, -OH, and -OCH ₃ ;

R ⁵ represents -H, or

and enantiomers, mixtures of enantiomers, diastereoisomers, mixtures of diastereoisomers, hydrates and solvates thereof.

Preferably, the compounds according to the present invention are selected from:

Saiginol B ^* R ⁵ = Rha (α/β),

Saiginol F,

Saiginol L,

Saiginol R. Please note that the abbreviation "Rha", as used herein, stands for the monosaccharide rhamnose. However, in preferred embodiments of the present invention "Rha", if not indicated otherwise, is L-rhamnose (also sometimes designated specifically as L-Rha). Even more preferably, the compounds according to the present invention are selected from:

Saiginol D,

Saiginol E,

Saiginol L,

Saiginol R. The inventive compounds according to general formula (I) accumulate in the floral petal and to a lesser extent in the stamen and pistil, but are absent in pollen (Fig. 1 ). However, inventors found that ColO populations do not produce the inventive compounds according to general formula (I). The presence or absence of the inventive compounds according to general formula (I) was assessed by LC-MS (Fig. 5).

Furthermore, inventors generated reciprocal introgression lines harboring chromosome segmental substitutions of ColO (non-producing the inventive compounds according to general formula (I)) in C24 (producing the inventive compounds according to general formula (I)), or substitutions of C24 in ColO were re-grown (see also Example 5). Intriguingly, inventors found a loss-of-f unction line in the C24 population which mapped to the same genome region as the gain-of-function line in the ColO population (see Fig. 4 and 6). Given that this genomic region on chromosome 2 of Arabidopsis harbors a total of 829 genes (Figure 4), the inventors further performed transcript profiling in order to compare gene expression in the gain-of-function donor line (C24) and its recurrent parent (ColO). Expression levels were generally very similar between the genotypes although a number of genes were significantly different between producing and non-producing accessions including several transposable elements.

However, only two genes were dramatically altered between ColO and C24, whereas the remaining 827 genes which were localized to the chromosomal segment substitution exhibited regular expression. The altered genes were renamed as putative flavonol Dhenylacyltransferases 1 (FPT1 - At2g22920) and 2 (FPT2 - At2g22960) (Fig. 7). The nucleic acid sequence of FPT2 (from C24) is referred to SED ID NO:1 and the nucleic acid sequence of FPT1 (from C24) is referred to SED ID NO:3.

Flavonol Phenylacyltransferase 2 (FPT2)

According to an aspect of some embodiments of the present invention a nucleic acid sequence, which has at least 80% sequence identity to SEQ-ID No.1 , wherein the nucleic acid sequence encodes a polypeptide having phenylacyltransferase activity, is provided. In other words one embodiment of the present invention refers to a nucleic acid sequence, which has at least 80% sequence identity to SEQ-ID No.1 , wherein the nucleic acid sequence encodes an enzyme having the same functionality as SEQ ID No. 1. Having the same functionality means that the enzyme or polypeptide comprises at least one part having phenylacyltransferase activity and thus is suitable to synthesize at least one compound of the invention. According to a preferred embodiment of the present invention, a nucleic acid sequence which has at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 81 %, at least about 82%, at least about 83%, at least about 84%, at least about 85%, at least about 86%, at least about 87%, at least about 88%, at least about 89%, at least about 90%, at least about 91 %, at least about 92%, at least about 93%, at least about 93%, at least about 94%, at least about 95%, at least about 96%, at least about 97%, at least about 98%, at least about 99%, or 100% sequence identity to SEQ-ID No.1 , wherein the nucleic acid sequence encodes a polypeptide having phenylacyltransferase activity, is provided.

The term "sequence identity", as used herein, indicates the percentage match of sequences by using an alignment between two sequences. An alignment is simply a correspondence between the sequences, in which each character in a sequence is assigned no more than one (maybe none) of the symbols in the other sequence, and in which the order of the symbols in the sequence is maintained, but in which gaps might be introduced into one or both sequence(s) to maximize identity. The sequence identity is usually and also in regard to the present invention calculated as the total number of matches (identical characters in both sequences at a certain position of the alignment) divided by the total length of the alignment of the two sequences and finally multiplied by 100. "Total length of the alignment", does thereby not refer to the length of the overlap between two aligned sequences, but to the entire length spanned by the aligned sequences, i.e. two sequences of 100 amino acids each, having an identical series of amino acids only in the last 10 amino acids of the first sequence and the first 10 amino acids of the second sequence, would result in a total alignment having a total length of 190 amino acids. Thus "sequence identity" is usually expressed as the percentage (%) of matches (identical characters) in positions from an alignment of two molecular sequences. The above concept can be used to determine the sequence identity of two polypeptide sequences (i.e. amino acid sequences) as well as of two nucleotide sequences (i.e. DNA or RNA). Sequences can be aligned with the use of a variety of computer programs known in the art, as for example BioEdit (Hall, T.A. (1999), Nucl. Acids. Symp. Ser. 41 , 95- 98). Another aspect of the present invention is directed to an isolated nucleic acid molecule comprising a nucleic acid sequence having at least 80% or higher (as mentioned before) sequence identity to SEQ-ID No. 1 , and encoding a polypeptide having phenylacyltransferase activity. Preferably no modifications like deletions, insertions or replacement of amino acids are made within the active site of the protein. Preferably all modifications which lead to the above-mentioned sequence identity deviant from 100% are present at amino acid positions of the enzyme outside the active site of the enzyme. In case one or more modifications are present within the active site of the enzyme, these modifications shall not decrease the catalytic activity of the enzyme. The present invention is also related to a nucleic acid molecule encoding a polypeptide which is capable of conferring resistance against UV-B irradiation in a plant, in which said polypeptide is expressed, said nucleic acid molecule comprising or consisting of a nucleic acid sequence encoding at least the mature form of a polypeptide comprising the amino acid sequence as given in SEQ-ID No. 2.

The present invention is also related to a nucleic acid molecule encoding a polypeptide which is capable of conferring resistance against UV-B irradiation in a plant, in which said polypeptide is expressed, said nucleic acid molecule comprising or consisting of a nucleic acid sequence comprising the coding regions of the DNA sequence as given in SEQ-ID No. 1. The present invention is also related to a nucleic acid molecule encoding a polypeptide which is capable of conferring resistance against UV-B irradiation in a plant, in which said polypeptide is expressed, said nucleic acid molecule comprising or consisting of a nucleic acid sequence encoding a polypeptide having at least 80% sequence identity to SEQ ID No: 2.

The present invention is also related to a nucleic acid molecule encoding a polypeptide which is capable of conferring resistance against UV-B irradiation in a plant, in which said polypeptide is expressed, said nucleic acid molecule comprising or consisting of a nucleic acid sequence encoding a polypeptide having at least 80%, at least 81 %, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ-ID No.2. Thereby it is preferred that the functionality or enzyme activity of said polypeptide is not changed or not decreased.

Nucleic acid molecules and nucleic acid sequences according to the present invention may be provided in recombinant form or free or substantially free of nucleic acid or genes of the species of interest or origin other than the sequence encoding a polypeptide with the required function. The nucleic acid molecules and nucleic acid sequences (and their encoded polypeptide products) may also be (i) isolated and/or purified from their natural environment (although not necessarily in pure form per se), or (ii) in substantially pure or homogeneous form.

Nucleic acid molecules and nucleic acid sequences according to the present invention may include cDNA, RNA, genomic DNA, preferably the intact gene, and may be wholly or partially synthetic (constructs'). Where a DNA sequence is specified, e. g. with reference to a figure or SEQ-ID No., unless context requires otherwise, the RNA equivalent, in which T (thymine) is replaced by U (uracil), is also encompassed. Also encompassed is the complement of the various disclosed sequences, which may be used in probing experiments, or in down-regulation of the sequence. Isolated nucleic acid molecules can be produced by standard techniques. For example, polymerase chain reaction (PCR) techniques can be used to obtain an isolated nucleic acid containing a nucleic acid sequence described herein. PCR can be used to amplify specific sequences from DNA as well as RNA, including sequences from total genomic DNA or total cellular RNA. Various PCR methods are described, for example, in PCR Primer: A Laboratory Manual, Dieffenbach and Dveksler, eds., Cold Spring Harbor Laboratory Press, 1995. Generally, sequence information from the ends of the region of interest or beyond is employed to design oligonucleotide primers that are identical or similar in sequence to opposite strands of the template to be amplified. Various PCR strategies also are available by which site-specific nucleic acid sequence modifications can be introduced into a template nucleic acid. Isolated nucleic acids can also be chemically synthesized, either as a single nucleic acid molecule (e.g., using automated DNA synthesis in the 3' to 5' direction using phosphoramidite technology) or as a series of oligonucleotides. For example, one or more pairs of long oligonucleotides (e.g., >100 nucleotides) can be synthesized that contain the desired sequence, with each pair containing a short segment of complementarity (e.g., about 15 nucleotides) such that a duplex is formed when the oligonucleotide pair is annealed. DNA polymerase is used to extend the oligonucleotides, resulting in a single, double-stranded nucleic acid molecule per oligonucleotide pair, which then can be ligated into a vector. Isolated nucleic acids of the invention also can be obtained by mutagenesis of, e.g., a naturally occurring DNA.

The term "isolated", as used herein, means that the material is removed from its original environment (e.g. the natural environment, if it is naturally occurring). For example, a naturally occurring nucleic acid present in a living plant is not isolated, but the same nucleic acid separated from some or all of the coexisting (e.g. proteins, other nucleic acids) materials in the natural system, is isolated. Such nucleic acids could be part of a vector and/or such nucleic acids could be part of a nucleic acid construct, and still be isolated in that such a vector or construct is not part of its natural environment. Nucleic acid molecules and nucleic acid sequences of the present invention may be optimized for plant expression. Examples of such sequence modifications include, but are not limited to, an altered G/C content to more closely approach the G/C content typically found in the plant species of interest, and the removal of codons atypically found in the plant species commonly referred to as codon optimization.

The phrase "codon optimization", as used herein, refers to the selection of appropriate DNA nucleotides for use within a structural gene or fragment thereof that approaches codon usage within the plant of interest. Therefore, an optimized gene or nucleic acid sequence refers to a gene in which the nucleic acid sequence of a native or naturally occurring gene has been modified in order to utilize statistically-preferred or statistically- favoured codons within the plant. The nucleic acid sequence typically is examined at the DNA level and the coding region optimized for expression in the plant species determined using any suitable procedure, for example as described in Sardana et al. (1996, Plant Cell Reports 15:677-681 ). A table of codon usage from highly expressed genes of dicotyledonous plants is compiled using the data of Murray et al. (1989, Nuc Acids Res. 17:477-498).

The nucleic acid sequence of the present invention can be, for example, utilized in preparation of recombinant polypeptides (or recombinant proteins) as well as in production of transformant plants with enhanced UV-B tolerance. In preparing recombinant polypeptides, generally, a nucleic acid sequence encoding the polypeptides of the present invention is inserted into an appropriate expression vector, the expression vector is transferred into an appropriate cell, the transformed cell is cultivated and the expressed polypeptides according to the invention is purified. According to an aspect of some embodiments of the present invention a polypeptide which has at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81 %, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ-ID No. 2, wherein the polypeptide has phenylacyltransferase activity, is provided.

Another aspect of the present invention is related to a nucleic acid sequence encoding for a polypeptide having at least at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81 %, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% sequence identity to SEQ-ID No. 2, wherein the polypeptide has phenylacyltransferase activity.

The present invention is also directed to an isolated nucleic acid molecule comprising a nucleotide sequence encoding a polypeptide having phenlyacyltransferase activity and having an amino acid sequence with at least 70% sequence identity to SEQ-ID No. 2. The present invention is also directed to an isolated nucleic acid molecule comprising a nucleotide sequence encoding a polypeptide having phenlyacyltransferase activity and having an amino acid sequence with at least 80% sequence identity to SEQ-ID No. 2.

The present invention is also directed to an isolated nucleic acid molecule comprising a nucleotide sequence encoding a polypeptide having phenlyacyltransferase activity and having an amino acid sequence with at least 90% sequence identity to SEQ-ID No. 2. The term "sequence identity", as used herein, indicates the percentage match of sequences by using an alignment between two sequences. An alignment is simply a correspondence between the sequences, in which each character in a sequence is assigned no more than one (maybe none) of the symbols in the other sequence, and in which the order of the symbols in the sequence is maintained, but in which gaps might be introduced into one or both sequence(s) to maximize identity. The sequence identity is usually calculated as the total number of matches (identical characters in both sequences at a certain position of the alignment) divided by the total length of the alignment of the two sequences and finally multiplied by 100. Thus "sequence identity" is usually expressed as the percentage (%) of matches (identical characters) in positions from an alignment of two molecular sequences. The above concept can be used to determine the sequence identify of two polypeptide sequences (i.e. amino acid sequences) as well as of two nucleotide sequences (i.e. DNA or RNA). Sequences can be aligned with the use of a variety of computer programs known in the art, as for example BioEdit (Hall, T.A. (1999), Nucl. Acids. Symp. Ser. 41 , 95-98).

As it was already mentioned above, the isolated nucleic acid molecule according to the present invention or the polypeptide according to the present invention shall not be modified in its sequence to an extent that diminishes or even abolishes the enzymatic function of the polypeptide according to the present invention or the polypeptide encoded by the nucleic acid molecule according to the present invention. This means the phenylacyltransferase activity shall not be diminished or abolished, since this would impair the capability to synthesize the inventive saiginols.

In this context, it is preferred, if the inventive polypeptide has at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81 %, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to SEQ-ID No. 2,

wherein within said percentage of sequence identity a sequence is included, which is 100% identical to a region (also termed "BLOCK 1 ") from amino acid position 58 to amino acid position 107, preferably from amino acid position 63 to amino acid position 102, more preferably from amino acid position 68 to amino acid position 97, more preferably from amino acid position 70 to amino acid position 90, even more preferably from amino acid position 73 to amino acid position 92, and even more preferably from amino acid position 78 to amino acid position 87 of SEQ-ID No. 2,

and wherein within said percentage of sequence identity also a sequence is included, which is 100% identical to a region (also termed "BLOCK 2") from amino acid position 155 to amino acid position 200, more preferably from amino acid position 160 to amino acid position 195, more preferably from amino acid position 165 to amino acid position 190, even more preferably from amino acid position 170 to amino acid position 185, and even more preferably from amino acid position 175 to amino acid position 180 of SEQ-ID No. 2.

It shall be understood that the given regions for BLOCK 1 and BLOCK 2 (as detailed above) can be freely combined with each other, i.e. it is not mandatory that the widest range of BLOCK 1 has to be combined with the widest range of BLOCK 2. Moreover, it shall be understood that the two regions, i.e. BLOCK 1 and BLOCK 2 can be separated from each other by a sequence of amino acids.

However, it is especially preferred in accordance with the invention, if the inventive polypeptide has at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81 %, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to SEQ-ID No. 2,

wherein within said percentage of sequence identity a sequence is included, which is 100% identical to a region (also termed "BLOCK 1 ") from amino acid position 78 to amino acid position 87 of SEQ-ID No. 2,

and wherein within said percentage of sequence identity also a sequence is included, which is 100% identical to a region (also termed "BLOCK 2") from amino acid position 175 to amino acid position 180 of SEQ-ID No. 2. It is even more preferred in accordance with the invention, if the inventive polypeptide has at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81 %, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to SEQ-ID No. 2,

wherein within said percentage of sequence identity a sequence is included, which is 100% identical to the amino acid sequence GPGCSSxxxL (SEQ-ID No. 21 ), wherein x represents a random amino acid,

and wherein within said percentage of sequence identity also a sequence is included, which is 100% identical to the amino acid sequence GDSYSG (SEQ-ID No. 22).

Concerning an inventive nucleic acid molecule, this is slightly more complicated, since the redundancy of the genetic code, i.e. several nucleotide triplets may code for more than one amino acid, allows a certain variation of the codon triplets without affecting the sequence of the final polypeptide. Thus, it is preferred, if the inventive nucleic acid sequence has at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81 %, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to SEQ-ID No. 1 ,

wherein within said percentage of sequence identity a sequence is included that encodes at least an amino acid sequence 100% identical to a region (also termed "BLOCK 1 ") from amino acid position 58 to amino acid position 107, preferably from amino acid position 63 to amino acid position 102, more preferably from amino acid position 68 to amino acid position 97, more preferably from amino acid position 70 to amino acid position 90, even more preferably from amino acid position 73 to amino acid position 92, and even more preferably from amino acid position 78 to amino acid position 87 of SEQ-ID No. 2, and wherein within said percentage of sequence identity also a sequence is included that encodes at least an amino acid sequence 100% identical to a region (also termed "BLOCK 2") from amino acid position 155 to amino acid position 200, more preferably from amino acid position 160 to amino acid position 195, more preferably from amino acid position 165 to amino acid position 190, even more preferably from amino acid position 170 to amino acid position 185, and even more preferably from amino acid position 175 to amino acid position 180 of SEQ-ID No. 2.

It is even more preferred, if the inventive nucleic acid sequence has at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81 %, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%sequence identity to SEQ- ID No. 1 ,

wherein within said percentage of sequence identity a sequence is included that encodes at least an amino acid sequence 100% identical to a region (also termed "BLOCK 1 ") from amino acid position 78 to amino acid position 87 of SEQ-ID No. 2,

and wherein within said percentage of sequence identity also a sequence is included that encodes at least an amino acid sequence 100% identical to a region (also termed "BLOCK 2") from amino acid position 175 to amino acid position 180 of SEQ-ID No. 2. Still even more preferred is an inventive nucleic acid sequence that has at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81 %, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99%, sequence identity to SEQ-ID No. 1 , wherein within said percentage of sequence identity a sequence is included that encodes at least the amino acid sequence GPGCSSxxxL (SEQ-ID No. 21 ), wherein x represents a random amino acid,

and wherein within said percentage of sequence identity also a sequence is included that encodes the amino acid sequence GDSYSG (SEQ-ID No. 22).

Amino acid or nucleotide positions, which are counted herein, start from the first letter in the respective sequence or SEQ-ID No. and start with "1 ". Transgenic plants

In order to experimentally test the function of the FPT proteins encoded by the DNA sequences SEQ-ID No. 1 and SEQ-ID No. 3, the inventors performed complementation assays in ColO and knockout fptl and fpt2 mutants. CaMV 35S driven complementation of the ColO fpt2 knockout mutant (Fig. 8) with the C24 allele of FPT2 resulted in the production of the compound according to the invention in flowers of the transgenic plant, however neither expression of the ColO allele of FPT2 in the ColO fpt2 knockout line nor expression of the ColO or the C24 allele in the ColO fptl mutants yielded flowers which produce compounds according to general formula (I). However, the total flavonol content is equivalent in all genotypes. Therefore, only FPT2 functions as a phenylacyltransferase in vivo, catalyzing the phenylacylation of precursors of compounds according to general formula (I) and accordingly finalizes the biosynthesis of compounds according to general formula (I).

Thus, another aspect of the present invention is related to convey the capability to produce the inventive saiginols to a plant or a plant cell.

Plant cells may be transformed stably or transiently with the nucleic acid constructs of the present invention. In stable transformation, the nucleic acid molecule of the present invention is integrated into the plant genome and as such it represents a stable and inherited trait. In transient transformation, the nucleic acid molecule is expressed by the cell transformed but it is not integrated into the genome and as such it represents a transient trait.

Techniques for introducing exogenous nucleic acid sequences to the genome of the chloroplasts are known to a person skilled in the art. For example, a technique for introducing exogenous nucleic acid sequences to the genome of the chloroplasts is involves the following procedures: First, plant cells are chemically treated so as to reduce the number of chloroplasts per cell to about one. Then, the exogenous nucleic acid is introduced via particle bombardment into the cells with the aim of introducing at least one exogenous nucleic acid molecule into the chloroplasts. The exogenous nucleic acid is selected such that it is integratable (i.e. it can be integrated) into the chloroplast's genome via homologous recombination, which is readily effected by enzymes inherent to the chloroplast.

Thus, a preferred embodiment of the present invention is directed to a method for expressing in the plant or plant cell a vector comprising a nucleic acid sequence having at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 81 %, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91 %, at least 92%, at least 93%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, or 100% identity to SEQ-ID No. 1.

The term "plant", as used herein, encompasses whole plants, ancestors and progeny of the plants and plant parts, including flower, leaf, root (including tubers), fruit, seed, shoot, stem, twig) and plant cells (homogeneous or heterogeneous populations of cells), tissues and organs. The plant may be in any form including suspension cultures, embryos, meristematic regions, callus tissue, leaves, gametophytes, sporophytes, pollen, and microspores. Plants that are particularly useful in the methods of the invention include all plants which are commercially or scientifically valuable and which taste, aroma or pigmentation modification is desired. A suitable plant for use with the method of the invention can be any potted plant, hydroponically-grown plant, field-grown plant, greenhouse-grown plant, in vitro-grown plant, in particular monocotyledonous and dicotyledonous plants including a fodder or forage legume, ornamental plant, food crop, tree, or shrub. According to an exemplary embodiment of the present invention, the plant is selected from the genus Arabidopsis.

However, transformation methods can be used for any plant, and are not limited to Arabidopsis. In general, the plants which can be modified according to the invention and which for example show overexpression of a polypeptide according to the invention can be derived from any desired plant species. They can be monocotyledonous plants or dicotyledonous plants, preferably they belong to plant species of interest in agriculture, wood culture or horticulture interest, such as crop plants (e.g. maize, rice, barley, wheat, rye, oats etc.), potatoes, oil producing plants (e.g. oilseed rape, sunflower, pea nut, soy bean, etc.), cotton, sugar beet, sugar cane, leguminous plants (e.g. beans, peas etc.), wood producing plants, preferably trees, etc. Following transformation of desired plant with the nucleic acid according to the invention or a nucleic acid sequence encoding the polypeptide according to the invention, a plant may be regenerated, e.g. from single cells, callus tissue or leaf discs, as is standard in the art. Almost any plant can be entirely regenerated from cells, tissues and organs of the plant. Even though both monocotyledonous plants and dicotyledonous plants can be used in accordance with the present invention (e.g. for transformation), it is especially preferred according to certain embodiments, if the plant used is dicotyledonous plant. In case the plant used in accordance with the invention is a dicotyledonous plant, it is especially preferred, if the plant is selected from Arabidopsis and/or tomato and/or tobacco.

In case the plant used in accordance with the invention is a monocotyledonous plant, it is preferred, if the plant is selected from maize (corn), rice, barley, wheat, rye, oats, sorghum, millet, triticale, fonio, teff, and wild rice; more preferably from maize (corn), rice, barley, wheat, rye, oats; and especially preferably the plant used in accordance with the invention is rice. Transient transformation can be effected by any of the direct DNA transfer methods described above or by viral infection using modified plant viruses.

In addition to the above, the nucleic acid molecule of the present invention can also be introduced into a chloroplast genome thereby enabling chloroplast expression.

According to an aspect of some embodiments of the present invention a vector comprising the nucleic acid sequence SEQ-ID No.1 is provided. Also preferred is a vector comprising the nucleic acid sequence according to claim 6. The present invention is also related to a vector comprising a nucleic acid sequence having at least 80% (or higher as disclosed herein) sequence identity to SEQ-ID No. 1.

The present invention is additionally related to a vector comprising a nucleic acid sequence having at least 80% sequence identity to SEQ-ID No. 1 , wherein the vector is an expression vector in which the nucleic acid sequence is operatively linked to one or more control sequences allowing the transcription and optionally expression in prokaryotic and/or eukaryotic host cells.

Another aspect of the present invention is related to a host cell containing a nucleic acid molecule comprising a nucleic acid sequence having at least 80% sequence identity (or higher as disclosed herein) to SEQ-ID No. 1 ;

or containing a vector comprising a nucleic acid sequence having at least 80% (or higher as disclosed herein) sequence identity to SEQ-ID No. 1 , wherein the vector is an expression vector in which the nucleic acid sequence is operatively linked to one or more control sequences allowing the transcription and optionally expression said host cell. The host cell according to the invention may be selected from bacterial cells, fungal cells, yeast cells, insect cells, vertebrate cells and plant cells. It is especially preferred in accordance with the invention, if the host cell is a plant cell. Thus, another aspect of the invention is related to a transgenic plant or a transgenic plant tissue comprising said plant cell.

"Vector", as used herein, is defined to include, inter alia, any plasmid, cosmid, phage or Agrobacterium binary vector in double or single stranded linear or circular form, which may or may not be self transmissible or mobilizable, and which can transform prokaryotic or eukaryotic hosts either by integration into the cellular genome or exist extrachromosomally (e.g. autonomous replicating plasmid with an origin of replication). Specifically included are shuttle vectors by which is meant a DNA vehicle capable, naturally or by design, of replication in two different host organisms, which may be selected from actinomycetes (and related species), bacteria and eukaryotic organisms (e.g. higher plant, mammalian, yeast or fungal cells). A vector including a nucleic acid according to the present invention does not need to include a promoter or other regulatory sequence, particularly if the vector is to be used to introduce the nucleic acid into cells for recombination into the genome.

Preferably the nucleic acid in the vector is under the control of, and operably linked to, an appropriate promoter or other regulatory elements for transcription in a host cell such as a microbial, e.g. bacterial, or plant cell. The vector may be a bifunctional expression vector which functions in multiple hosts. In the case of genomic DNA, this may contain its own promoter or other regulatory elements and in the case of cDNA this may be under the control of an appropriate promoter or other regulatory elements for expression in the host cell.

By "promoter", as used herein, is meant a sequence of nucleotides from which transcription may be initiated of DNA operably linked downstream (i.e. in the 3-direction on the sense strand of double-stranded DNA).

"Operably linked", as used herein, means joined as part of the same nucleic acid molecule, suitably positioned and oriented for transcription to be initiated from the promoter. DNA operably linked to a promoter is "under transcriptional initiation regulation" of the promoter. Another embodiment of this aspect of the present invention provides a gene construct, preferably a replicable vector, comprising an inducible promoter operatively linked to a nucleotide sequence provided by the present invention.

The term "inducible" as used in connection with "promoter" is well understood by those skilled in the art. In essence, expression under the control of an inducible promoter is "switched on" or increased in response to an applied stimulus. The nature of the stimulus varies between promoters. Some inducible promoters cause little or undetectable levels of expression (or no expression) in the absence of the appropriate stimulus. Other inducible promoters cause detectable constitutive expression in the absence of the stimulus. Whatever the level of expression is in the absence of the stimulus, expression from any inducible promoter is increased in the presence of the correct stimulus. The preferable situation is where the level of expression increases upon application of the relevant stimulus by an amount effective to alter a phenotypic characteristic. Thus an inducible (or "switchable") promoter may be used which causes a basic level of expression in the absence of the stimulus which level is too low to bring about a desired phenotype (and may in fact be zero). Upon application of the stimulus, expression is increased (or switched on) to a level which brings about the desired phenotype.

Possible regulatory elements permitting expression in prokaryotic host cells comprise, e.g., the PL, lac, trp or tac promoter in E. coli, and examples for regulatory elements permitting expression in eukaryotic host cells are the AOX1 or GAL1 promoter in yeast or the CMV-, SV40- , RSV-promoter (Rous sarcoma virus), CMV- enhancer, SV40-enhancer or a globin intron in mammalian and other animal cells. In this context, suitable expression vectors are known in the art such as Okayama-Berg cDNA expression vector pcDV1 (Pharmacia), pCDM8, pRc/CMV, pcDNAI , pcDNA3 (Invitrogen), pSPORTI (GIBCO BRL).

Particularly of interest in the present context are plant vectors. Specific procedures and vectors previously used with wide success upon plants are described in the prior art. Suitable promoters which operate in plants include the Cauliflower Mosaic Virus 35S (CaMV 35S) gene promoter that is expressed at a high level in virtually all plant tissues, the cauliflower meri 5 promoter that is expressed in the vegetative apical meristem as well as several well localised positions in the plant body, e.g. inner phloem, flower primordia, branching points in root and shoot and the Arabidopsis thaliana LEAFY promoter that is expressed very early in flower development. Other promoters include the rice actin promoter, tomato fruit-specific E8 promoter and flower specific promoter APETALA3 are suitable for the purpose.

The promoter may include one or more sequence motifs or elements conferring developmental and/or tissue specific regulatory control of expression.

Thus the vectors according to the invention may comprise a nucleotide sequence having at least 80% sequence identity (or higher as disclosed herein) to SEQ-ID No. 1 , in addition to various sequences required to give them replicative, integrative and/or expression functionality. Such vectors can be used, for instance, to make plants into which they are introduced resistant to UV-B irradiation. According to an aspect of some embodiments of the present invention a nucleic acid construct comprising a promoter that functions in plant cells is provided, operably linked to the nucleic acid sequence SEQ-ID No.1.

Conferring UV-B tolerance of FPT2 and by implication of compounds according to general formula (I) in plants was surprisingly proven by achieving an increased seed yield of FPT2- C24 overexpressing lines (FPT2C24-OX - Fig. 9b) and the gain-of-function NIL N09, in comparison to the non-producing wildtype ColO as well as to the flavonoid less tt4 mutant lines, while being subjected to a range of UV-B treatments. The vigor of the NIL N09 and FPT2-C24 overexpressing lines was reflected in the number of leaves and inflorescences these lines produced (Fig. 9a).

The seed yield was considerably higher in the producing line (FPT2C24-OX) and the gain- of-function line (NIL N09) but also the germination rate of the seeds was doubled (Fig. 9c), revealing that the production of germinable seeds was almost five times higher in the gain- of-function (NIL N09) line and the C24-FPT2 overexpressing line than in lines that do not produce compounds according to general formula (I) (Fig. 9c). Thus, another aspect of the present invention is related to a method of conferring UV-B tolerance to a plant, wherein the method comprises the steps of:

(i) transforming a recipient plant with a vector comprising the nucleic acid sequence SEQ-ID No.1 ;

(ii) producing one or more transgenic offspring of the recipient plant; and

(iii) selecting the offspring for UV-B tolerance.

Thus, another aspect of the present invention is related to a method of conferring UV-B tolerance to a plant, wherein the method comprises the steps of:

(i) transforming a recipient plant with a vector comprising a nucleic acid sequence having at least 80% sequence identity to SEQ-ID No.1 ;

(ii) producing one or more transgenic offspring of the recipient plant; and

(iii) selecting the offspring for UV-B tolerance.

Thus, another aspect of the present invention is related to a method of conferring UV-B tolerance to a plant, wherein the method comprises the steps of:

(i) transforming a recipient plant with the nucleic acid according to claim 6 or 7 or the vector according to claim 8 or 9;

(ii) producing one or more transgenic offspring of the recipient plant; and

(iii) selecting the offspring for UV-B tolerance. The sequence identity may be higher than 80% as disclosed herein so that any other of the sequence identity values as discloses herein can be used instead of the 80%.

The particular choice of a transformation technology will be determined by its efficiency to transform certain plant species as well as the experience and preference of the person practicing the invention with a particular methodology of choice. It will be apparent to the skilled person that the particular choice of a transformation system to introduce nucleic acid according to the invention into plant cells is not essential to or a limitation of the invention, nor is the choice of technique for plant regeneration.

As used herein, the phrase "UV-B tolerance" refers to genetically modified organisms, withstanding UV-B irradiation without damage of RNA, DNA or proteins for a period of time, compared to wildtype organisms. Thus, "UV-B tolerance" can for example be assessed by measuring vital parameters, known to a person skilled in the art, of organisms after UV-B irradiation treatment for 2 h per day for 28 days with 1000 mW/m ² .

As used herein, the phrase "UV-B tolerant plant" refers to genetically modified plants, withstanding UV-B irradiation without damage of RNA, DNA or proteins for a period of time, compared to wild type plants. Thus, "UV-B tolerance" of a plant can for example be assessed by measuring vital parameters, known to a person skilled in the art, of plants after UV-B irradiation treatment for 2 h per day for 28 days with 1000 mW/m ² .

Another aspect of the present invention is directed to a method for producing the compounds of general formula (I) - (IV) comprising expressing, overexpressing or synthesizing products of one or more nucleic acid molecules according to the invention in a host cell.

Therefore one preferred embodiment of the invention refers to a method for the preparation of at least one compound according to anyone of the general formulae (I) - (IV) using a UV-B tolerant plant comprising a nucleic acid that encodes a polypeptide which is suitable to synthesize a compound of anyone of the general formulae (I) - (IV) and wherein said nucleic acid has been introduced into the UV-B tolerant plant by genetic manipulation. In other words one preferred embodiment of the invention refers to a method for preparation of at least one compound according to anyone of the general formulae (I) - (IV) using a transgenic UV-B tolerant plant, wherein a nucleic acid which has been introduced into the UV-B tolerant transgenic plant codes for an enzyme suitable for synthesizing of at least one compound according to anyone of the general formulae (I) - (IV), preferably coding for FPT2. In general a host, preferably a plant such as a tobacco plant or an Arabidopsis line, is transformed to introduce a nucleic acid construct coding for the enzyme FPT2, such as a construct comprising the SEQ ID No. 1 and a suitable promotor. The introduced nucleic acid construct results in a transgenic plant, which overexpresses the enzyme FPT2 or which expresses the enzyme FPT2 only because of the introduced (transformed) nucleic acid construct.

The present invention is also directed to a method for producing the compounds of any one of the general formulae (I) - (IV) comprising expressing, overexpressing or synthesizing products of one or more nucleic acid molecules according to the invention in a host cell, wherein the host cell is a plant cell.

According to an aspect of some embodiments of the present invention a transgenic or genetically modified UV-B tolerant plant comprising a vector according to the present invention is provided, wherein the UV-B tolerant plant synthesizes the inventive compounds of anyone of the general formulae (I) - (IV). This means the plant has an improved UV-B tolerance because of introduction of the nucleic acid construct coding for the flavonol phenylacyltransferase by genetic manipulation, such as transformation. Therefore, one embodiment of the present invention is a UV-B tolerant transgenic plant comprising the nucleic acid sequence according to claim 6 or 7 or the vector according to claim 8 or 9 that synthesizes a compound of anyone of the general formulae (I) - (IV) and wherein the nucleic acid according to claim 6 or 7 or the vector according to claim 8 or 9 has been introduced into the UV-B tolerant transgenic plant by genetic manipulation. It is thereby preferred that the UV-B tolerant plant of the invention contains a vector comprising sequence ID No.1. Methods for transformation of plants, such as gene gun or viral infection, are commonly known in the art.

According to an aspect of some embodiments of the present invention an UV-B tolerant plant comprising a vector according to the present invention is provided, wherein the UV-B tolerant plant displays enhanced synthesis of the inventive compounds of anyone of the general formulae (I) - (IV).

As used herein, the term "enhanced" or "enhancing" refers to an increase in production of a compound according to anyone of the general formulae (I) - (IV), for example, by at least 5%, 10%, 15%, 20%, 30%, 50%, 100%, 200%, 250%, 400% or more. The increase in production of a compound according to general formula (I) may be in the emission level or in the internal pool level and can typically be determined with respect to a native plant (i.e., a plant not modified with the biomolecules of the invention, i.e. a non-genetically modified plant of the same species which is grown under the same growth conditions). The enhanced production of a compound according to anyone of the general formulae (I) - (IV) may be determined by any method known to one of ordinary skill in the art, as for example by headspace sampling, solvent extraction, solid phase micro extraction and the analysis by gas chromatography-mass spectrometry or any other analytical instrument. According to an aspect of some embodiments of the present invention a transgenic or genetically modified UV-B tolerant plant comprising a nucleic acid construct according to the invention is provided, wherein the UV-B tolerant plant or the UV-B tolerant transgenic plant synthesizes at least one inventive compound of one of the general formulae (I) - (IV).

According to an aspect of some embodiments of the present invention a transgenic or genetically modified UV-B tolerant plant comprising a nucleic acid construct according to the invention is provided, wherein the UV-B tolerant plant display synthesis or enhanced synthesis of at least one inventive compound of anyone of the general formulae (I) - (IV). Thereby the synthesis or enhanced synthesis is caused by a genetic manipulation. It is preferred that this genetic manipulation introduces said nucleic acid construct. This means the synthesis of at least one inventive compound of anyone of the general formulae (I) - (IV) is increased compared to the wildtype plant without said nucleic acid construct of the present invention.

Thus, a further aspect of the present invention is directed to a UV-B tolerant plant comprising the nucleic acid according to claim 6 or 7 or the vector according to claim 8 or 9 that synthesizes a compound according to anyone of the general formulae (I) - (IV) and wherein the nucleic acid according to claim 6 or 7 or the vector according to claim 8 or 9 has been introduced into the UV-B tolerant plant by genetic manipulation.

Another aspect of the present invention relates to a UV-B tolerant plant comprising the nucleic acid construct according to claim 12 that synthesizes a compound according to anyone of the general formulae (I) - (IV), wherein said nucleic acid construct has been introduced into the UV-B tolerant plant by genetic manipulation.

In another embodiment of the present invention purified or isolated nucleic acids or nucleic acid fragments complementary or antisense to the nucleic acids or nucleic acid fragments encoding FPT2 or functionally active fragments or variants thereof are provided. The present invention also provides substantially purified or isolated nucleic acids or nucleic acid fragments encoding amino acid sequences for a class of proteins which are related to FPT2, or functionally active fragments or variants thereof. Such proteins are herein referred to as FPT2-like.

By a FPT2-like polypeptide is meant that either one or both of the following criteria apply: (i) the gene which encodes the polypeptide is expressed in a similar manner to FPT2, and (ii) the polypeptide has similar functional activity to FPT2.

Use of the saiginols according to the invention

According to an aspect of some embodiments of the present invention a composition comprising the compound according to anyone of the general formulae (I) - (IV) is provided, optionally containing a cosmetically-acceptable carrier or excipient. According to an aspect of some embodiments of the present invention a composition containing an inventive compound according to anyone of the general formulae (I) - (IV) for use as sunscreen is provided. According to an aspect of some embodiments of the present invention a composition containing an inventive compound according to anyone of the general formulae (I) - (IV) for protection from UV-B radiation is provided.

According to an aspect of some embodiments of the present invention compounds according to anyone of the general formulae (I) - (IV) for use as UV-B protectants is provided.

According to an aspect of some embodiments of the present invention compounds according to anyone of the general formulae (I) - (IV) for use as UV-B protectants in a sunscreen is provided.

Another aspect of the present invention is the use of the compounds according to anyone of the general formulae (I) - (IV) in a sunscreen composition. Further provided is the cosmetic use of compounds according to the present invention as UV-B protectants.

The concentration of the compounds according to anyone of the general formulae (I) - (IV) in a composition according to the invention may be from about 0.1 % by weight to about 10% by weight, preferably from about 0.5% by weight to about 5% by weight, preferably from about 0.5% by weight to about 2% by weight.

Compositions of the present invention may include a volatile solvent. By "volatile", as used herein, it is meant compounds that meet one or more of the following requirements: those compounds with a flash point below 150°C, such as less than about 130°C, such as less than about 50°C, such as less than about 25°C, and/or a boiling point of less than about 150°C, such as less than about 100°C, such as less than about 90°C.

By "solvent" it is meant, a compound that is capable (absent any other ingredients such as surfactants, co-solvents, etc.) of dissolving (e.g., forming a clear solution, such as one that transmits at least 25%, such as at least about 50% of light intensity of a 700nm wavelength source through a 1 cm path length, as measured by conventional visible spectrophotometry) the compounds according to general formula (I). The volatile solvent is preferably cosmetically-acceptable and is preferably a liquid at room temperature. Furthermore, the volatile solvent is, in certain embodiments, miscible with water. By "miscible with water", as used herein, it is meant that the volatile solvent is fully soluble in water in all proportions. Suitable examples of volatile solvents that are miscible with water include ethanol, propanol, and isopropanol. The volatile solvent, if not miscible with water, may, in certain embodiments, be soluble to at least some extent in isopropanol.

The amount of volatile solvent present in the composition may be from 10% to 60%, or from 20% to 55%, or from 30% to 50%. In certain embodiments, it is preferred if titanium oxide is present in the composition from 10% to 25%.

The composition of the invention can also comprise active materials and/or beneficial agents, such as conditioning agents, moisturising agents, emollients, astringent or antiperspirant compounds, biocidal compounds, sunscreens or UV absorbers, pigments, perfumes, anti-aging agents, enzymes, proteins, and vitamins.

As moisturizing agents can be mentioned for example glycerol, sorbitol, urea, collagen, gelatine, aloe vera, and hyaluronic acid. They can represent up to about 10% of the total weight of the composition.

In embodiment, where at least one emollient is present, the at least one emollient is preferably selected from alkylmonoglycerides, alkyldiglycerides, triglycerides such as oils extracted from plants and from vegetables (e.g., palm oil, coprah oil, cottonseed oil, soya bean oil, sunflower seed oil, olive oil, grapeseed oil, sesame oil, peanut oil, castor oil) or oils of animal origin (e.g., tallow, fish oils, etc.), derivatives of these oils such as hydrogenated oils, lanolin derivatives, mineral oils or paraffin oils, perhydrosqualane, squalene, diols such as 1 ,2-propanediol, 1 ,3-butanediol, cetyl alcohol (also referred to as 1 -hexadecanol), stearyl alcohol, oleic alcohol, polyethylene glycols or polypropylene glycols, fatty esters such as isopropyl palmitate, 2-ethylhexyl cocoate, myristyl myristate, esters of lactic acid, stearic acid, behenic acid, isostearic acid, silicone oils such as polydimethylsiloxanes, silicone copolyols (dimethicone copolyol, cetyldimethicone copolyol), diphenyl-dimethicones, phenyltrimethicones, dimethiconols, with viscosities in the range about 20 to about 10000 mPa ^* s. They can represent up to about 20% by weight of the composition.

Flavour, essential oils, perfumes can for example include benzaldehyde, caraway oil, cardamon oil, cinnamon oil, ethylvanilin, eucalyptus globulus oil, glutamic acid, clove oil, orange oil, peppermint oil, thymol, phenethyl alcohol or their mixtures. They can represent up to about 3% of the total weight of the composition.

Proteins can for example include collagen, collagen derivatives and keratin. They can represent up to about 3% of the total weight of the composition. Retinol, retinyl palmitate, tocopherol, tocopherol acetate, menadione, ascorbic acid and ascorbyl palmitate are examples of vitamins that can be included in the present compositions. They can represent up to about 0.5% of the total weight of the composition. In another embodiment of the composition according to the invention said composition is in form of a cream, lotion, liquid or gel, with the effectiveness of a sunscreen.

Another aspect of the present invention is a composition for use as sunscreen comprising: a. Compounds according to general formula (I) in an amount from 0.1 % to 10% of the total weight of the composition

b. A volatile solvent in an amount of 10% to 60% of the total weight of the composition c. Titanium oxide in an amount from 10% to 25% of the total weight of the composition d. Moisturizing agents in an amount up to 10% of the total weight of the composition e. Emollients in an amount up to 20% of the total weight of the composition

f. Proteins in an amount up to 3% of the total weight of the composition

g. Flavour in an amount up to 3% of the total weight of the composition

h. Vitamins in an amount up to 0.5% of the total weight of the composition It is clear to a skilled person that all ingredients of the sunscreen shall add up to 100% by weight.

An exemplary sunscreen composition containing the following components can be prepared according to the invention. Such a sunscreen may contain as active ingredients 15% - 25% by weight titanium oxide and 5.0% - 10.0% by weight saiginol A and may contain as inactive ingredients 50% - 70% by weight ethanol, 2.0% - 3.0% by weight hyaluronic acid, 3.0% - 7.0% by weight grape seed oil, 1.5% - 3.5% by weight sesame oil, 1.5% - 3.5% by weight sunflower seed oil, and 0.05% - 0.15% by weight tetrahexyldecyl ascorbate.

A quality of the instant composition is that the compounds according to general formula (I) have a critical wavelength of about 280 nm or greater.

In another embodiment, the sunscreen composition according to the invention has an SPF of at least 20, preferably an SPF of at least 25, more preferably SPF of at least 30, an even more preferably an SPF of at least 35.

The abbreviation "SPF", as used herein, stands for "sunburn protection factor" (sometimes also referred to as "sun protection factor"). The SPF is a measure of how much solar energy (in particular by UV radiation) is required to produce sunburn on protected skin (i.e. in the presence of sunscreen) relative to the amount of solar energy required to produce sunburn on unprotected skin (i.e. without sunscreen). An increasing SPF value means that also sunburn protection increases. The SPF can be determined experimentally under reproducible conditions on a meaningful number of test subjects (see for example ISO 24444:2010).

In a preferred embodiment of the invention the composition can be used as cosmetics with sunscreen effect. Description of the figures

Tissue specificity of the accumulation of saiginols. The relative ratio was estimated by using total peak area of general flavonol-glycosides and phenylacylated flavonol-glycosides.

Structural analysis of saiginol. A computational estimation of the most stable stereochemical structure of saiginol A is shown. The estimation was performed by MMFF94 of Marvin (http://www.chemaxon.com/).

Spectrometric analysis of saiginol. HPLC-PDA profiling of a) kaempferol- 3-0-Glc-2"-0-Rha-7-0-Rha, b) saiginol A (kaempferol-3-0-Glc-2"-0-Rha-6"- sinapoyl-7-O-Rha) and c) sinapoyl-GIc in the spectral range between 260nm and 450nm.

Chromosomal mapping of metabolite Quantitative Trait Loci (mQTL) for saiginol production. The chromosomal region on chromosome 2 indentified to be responsible for saiginol production (At2g22230-At2g3160) is delimited by a frame.

Saiginol A production in near isogenic lines (NIL) of ColO and C24 accessions. Chromatograms of gain-of-function and loss-of-function near isogenic lines (NIL) obtained from mQTL analysis of reciprocal crosses between ColO and C24 are shown. N and M lines are ColO and C24 background introgression lines, respectively. Chromatograms show ion extracted chromatograms for saiginol A (945m/z), B & I (901 m/z), C (885m/z) and D (799m/z), respectively. mQTL analysis of saiginol A in near isogenic lines (NIL) of ColO and C24 accessions. In total 45 M lines (C24 background) and 69 N lines (ColO background) were used for saiginol targeted mQTL analysis. The heatmap shows values of saiginol A production displaying on a log ₂ scale divided by the average of all values.

Fig. 7: shows a microarray analysis of gene expression of the genes encoded in the genomic region of chromosome 2 encoding the mQTL in five producing accessions (C24, Cvi, Da, Rsch and RLD) and five non-producing accessions (ColO, La-er, Ws, Sap and Stw). Duplicate hybridisations were carried out for ColO and C24 for all other accessions a single hybridization was performed. Intensity indicates fold change compared to the expression level in ColO.

In vivo functional characterization of FPT1 and 2 by metabolite profiling of transgenic plants.

All transgenic lines shown have a ColO background. FPU (ColO), FPT1 (C24), FPT2-1 (ColO), FPT2-2(ColO) and FPT2(C24) indicate FPT1 cloned from ColO and C24, FPT2 cloned from ColO (FPT2-1 is shortest transcript and FPT2-1 is longest transcript) and C24 respectively. The chromatograms show ion extracted chromatogram for saiginol A (945m/z), B & I (901 m/z), D (799m/z), G (961 m/z) and J (815m/z), respectively.

Functional characterization of the role of saiginols in protection against UV-B irradiation.

a) Representative photographs of the phenotypic changes of genotypes exhibiting modified flavonol contents following UV-B irradiation treatment (2h per day for 28 days with 1000 mW/m2 UV-B light irradiation)

b) Seed yield after long-term (2 h per day for 28 days) UV-B treatment (1000 and 2000 mW/m2). Seeds were collected on a single plant basis but three individual plants per genotype were measured. Data is presented as mean ± SE. *P<0.05.

c) Germination rate using seeds obtained after UV-B irradiation (2h per day for 28 days with 2000 mW/m ² UV-B light irradiation).

Rate of silique production under UV-B irradiation after long-term (2 h per day) UV-B treatment (1 W/m ² ) using the detached immature inflorescences of first bolting. Error bars indicate the SE of six biological replicates. Data is presented as mean ± S.E.M. *P<0.05. Abbreviation foe indication of plant lines is as written in example 10. OX-1 and OX-2 designate independent overexpressing lines using the same construct.

AtFPT2 overexpressing transgenic tobbaco plants.

Three metabolites (805, 821 , 835 m/z), which are annotated phenyacylated flavonol-glycosides were observed in AtFPT2-OX tobacco transgenic plants. This figure shows results of example 13. EXAMPLES

Plant material

Accessions of Arabidopsis thaliana used in the present application were obtained from NASC the European Arabidopsis Stock Centre (http://arabidopsis.info/). Plants were cultured on agar plates in a growth chamber under normal long day light conditions (16 h day, 140-160 μηηοΙ photons m ^"2 s ^"1 , 20°C; 8 h night, 16°C) for 14 days and transferred to soil. Flower materials were harvested from individual plants, immediately frozen in liquid nitrogen, and stored at -80°C until further use.

Example 1 : Secondary metabolite profiling by LC-MS

Given that considerable evidence has accumulated concerning the UV-B protective function of various phenylpropanoids during the processes of flower development, pollination and seed production assessment of levels of various phenylpropanoids in flowers in a set of 72 Arabidopsis ecotypes, which provided good coverage of the overall natural variability of the species, was performed by the inventors. Application of liquid chromatography-mass spectrometry (LC-MS) resulted in the detection of a total of 67 peaks consisting of 21 flavonoids, 12 glucosinolates, five hydroxycinnamates, eight polyamines, three putative lipid derivatives and 18 peaks of unknown chemical structure. All data were processed using Xcalibur 2.1 software (Thermo Fisher Scientific, Waltham, USA). Peak identification and annotation were performed in a combination approach using standard chemical confirmation, MSMS profiling, retention time profiling, mutant analysis, literature survey. In order to carry out mutant analysis for flavonoid derivatives, 14 mutants; ugt78d2 mutant (flavonoid-3-O-glucoside-less), tt7 mutant (quercetin and isorhamnetin derivatives-less), ugt78d1 mutant (flavonol-3-O-rhamnosides-less), ugt78d3 mutant (flavonol-3-O-arabinosides-less), omtl mutant (isorhamnetin-derivatives-less), ugt89c1 mutant (flavonol-7-O-rhamnosides-less), tt4 mutant (all flavonoid-less), and papl- D mutant (anthocyanin-overaccumulator), and La-er background tt mutant series obtained from NASC (tt3 , tt4, tt5, tt6) were used.

Example 2: Procedure of purification and characterization of saiginol A

MS-MS fragmentation studies suggested that these 18 peaks of unknown identity from Example 1 are novel flavonol derivatives. In order to characterize the chemical structure of these peaks, large amounts of the Arabidopsis accession C24, which accumulated these compounds, were re-grown and processed. Column chromatography was carried out over ODS (Nacalai Tesque, Cosmosil 75C18- OPN). HPLC analysis was carried out on an Atlantis ^® (φ 4.6 χ 150 mm, Waters) at a flow rate of 0.5 ml min ^"1 . Preparative HPLC was performed on a LC 10A system (Shimadzu) using an Inertsil ^® ODS-EP 5 μηη (φ 6.0 χ 150 mm) at 30 °C and monitoring was accomplished by PDA (200-600 nm). HPLC-PDA-ESI-MS (HPLC/photodiode array detection/electrospray ionization mass spectrometry) was performed on a Finnigan LCQ- DECA mass spectrometer (ThermoQuest, San Jose, CA, USA) and an Agilent HPLC 1100 series (Agilent Technologies, Palo Alto, CA, USA). HR-ESI-MS was performed on an ExactiveTM mass spectrometer (ThermoQuest, San Jose, CA, USA). Optical rotations were determined on a JASCO P-1020. UV spectra were recorded on a JASCO V-560. NMR data were recorded on JEOL JNM ECP-600. The deuterated solvent CD3OD was used for peak 4. Coupling constants are expressed in Hz. Plant materials, extraction and purification

Plant samples (26.72 g fresh weight) were harvested and immediately frozen in liquid nitrogen, the whole of which was immediately extracted with methanol. After concentration, MeOH liquid extraction was extracted with n-hexane, CHCI3 to remove low polarity metabolites. After the liquid-liquid partition and concentration, MeOH soluble fraction was obtained and was dissolved with H ₂ O. After liquid-liquid partition with n- BuOH, the n-BuOH fraction was obtained (256.9 mg).

This fraction was applied to ODS column (φ 3.5 χ 7 cm), and roughly separated by eluting with a gradient of H ₂ O as solvent A and CH ₃ CN as solvent B and the following elution profile [fraction 1 : 0% CH ₃ CN, fraction 2: 10% CH ₃ CN, fraction 3: 20% CH ₃ CN, fraction 4: 30% CH ₃ CN, fraction 5: 40% CH ₃ CN, fraction 6: 50% CH ₃ CN, fraction 7: 60% CH ₃ CN, fraction 8: 70% CH ₃ CN, fraction 9 80% CH ₃ CN and fraction 10: 100% CH ₃ CN (elution solvent: 70 ml/fraction)] to give 10 fractions. After LC-MS analysis for trace of peak 4, fraction 2 to 4 was assembled (fraction A). Fraction A was applied to ODS column (φ 3.5 x 7 cm) again, and separated by eluting with a gradient of H ₂ O as solvent A and CH ₃ CN as solvent B and the following elution profile [fraction A-1 and A-2: 0% CH ₃ CN, fraction A-3 and A-4: 10% CH ₃ CN, fraction A-5 to A-7: 20% CH ₃ CN, fraction A-8 to A-11 : 30% CH ₃ CN and fraction A-12: 100% CH ₃ CN (elution solvent: 30 ml (fraction A-1 to A-5), 15 ml (fraction A-6 to A-12) to give 12 fractions.

After LC-MS analysis for trace of peak 4, fraction A-10 and A-11 were assembled (fraction B). Fraction B (12.0 mg) was applied to preparative HPLC using an isocratic elution (20% CH ₃ CN in H ₂ O) at a flow rate of 4 ml/min to give peak 4 (3.5 mg). Characterization of saiginol A

Kaennpferol-3-0-[2-0-(a-L-rhannnopyranosyl)-6-0-(sinapoyl)- -D-glucopyranoside]-7-0-a- L-rhamnopyranoside

Yellow amorphous solid. [a] _D ²⁰ - 54.1 ° (c 0.09, MeOH). UV (MeOH): X _max (logs): 201.5 (3.47), 223.0 (3.61 ), 266.5 (3.37), 331.5 (3.47). HRESI-MS: m/z 969.2637 ([M+Na] ⁺ , calcd. for C ₄₄ H ₅₀ O ₂ 3Na, 969.2635). ¹ H NMR (CD ₃ OD) δ: 6.25 (1 H, br.s, H-6), 6.41 (1 H, br.s, H-8), 7.90 (2H, d, J = 8.8 Hz, H-2' and H-6'), 6.85 (2H, d, J = 8.8 Hz, H-3' and H-5') (kaempferol; H-6, 8, H-2' to 6'), 5.75 (1 H, d, J = 7.2 Hz, H-1 ), 3.63 (1 H, m, H-2), 3.58 (1 H, m, H-3), 3.32 (1 H, m, H-4), 3.51 (1 H, m, H-5), 4.26 (1 H, m, H-6a), 4.40 (1 H, m, H-6b) (glucose; H-1 to H- 6a,b), 5.23 (1 H, br.s, H-1 ), 3.99 (1 H, m, H-2), 3.75 (1 H, m, H-3), 3.32 (1 H, m, H-4), 3.97 (1 H, m, H-5), 0.90 (1 H, m, H-6) (rhamnose 1 ; H-1 to H-6), 6.66 (2H, s, H-2), 6.06 (1 H, d, J= 16.0, H-7a), 7.28 (1 H, d, J= 16.0, Η-8β), 3.87 (6H, s, -OCH ₃ ) (sinapic acid; H-2, H-7a, Η-8β and -OCH ₃ ), 5.33 (1 H, br.s, H-1 ), 3.47 (1 H, m, H-2), 4.04 (1 H, m, H-3), 3.76 (1 H, m, H-4), 3.47 (1 H, m, H-5), 1.18 (1 H, m, H-6) (rhamnose 2; H-1 to H-6). ¹³ C NMR (CD ₃ OD) δ: 161.7, 134.5, 179.7, 159.7, 99.9, 163.2, 96.4, 157.9, 107.5, 123.2, 132.5, 116.4, 163.0, 116.4, 132.5 (kaempferol; C-2 to C-10 and C-1 ' to C-6'); 100.2, 80.2, 79.2, 74.3, 76.1 , 64.6 (glucose; C-1 to C-6); 102.8, 70.4, 72.5, 72.6, 72.6, 17.2 (rhamnose 1 ; C-1 to C-6), 12.75, 107.0, 148.5, 139.5, 149.5, 107.0, 115.6, 147.0, 168.7 (sinapic acid; C1- to C-9), 99.9, 71.4, 71.8, 72.3, 73.9, 19.3 (rhamnose 2; C-1 to C-6). HMBC correlations from the anomeric proton (Rha1 H-1 , δ 5.23) to C-2 of glucose, the protons (glucose H-6a,b, δ 4.26 and 4.40) to C-9 of sinapic acid and the anomeric proton (Rha2 H-1 , δ 5.33) to C-7 of kaempferol were observed.

Example 3: UV-B absorption profile of Saiginol A

UV-VIS spectra in the range between 260nm and 450nm were recorded and analyzed via HPLC-PDA profiling for kaempferol-3-Glc-2"-Rha-7Rha (Fig. 3A), saiginol A (kaempferol- 3-Glc-2"-Rha-6"-sinapoyl-7Rha, Fig. 3B) and sinapoyl-glucose (Fig. 3C). HPLC-PDA (HPLC/photodiode array detection) was performed on Dionex Ultimate 3000 system (San Jose, CA, USA). Column chromatography was carried out on a Luna ^® (φ 2.00 χ 150 mm, phenomenex) at a flow rate of 0.2 ml min ^"1 and monitoring was accomplished by PDA (200-600 nm).

The flavonol-glycoside kaempferol-3-Glc-2"-Rha-7Rha, a precursor of saiginol A, appeared to be much less efficient in UV-B absorption than the phenylacylated saiginol A (kaempferol-3-Glc-2"-Rha-6"-sinapoyl-7Rha)(see Fig. 3A, B). The UV-VIS spectra of a Saiginol A revealed a Amax at 201.5 (3.47), 223.0 (3.61 ), 266.5 (3.37) and 331.5nm (3.47).

The substances kaempferol-3-0-Glc-2"-0-Rha-7-0-Rha and sinapoyl-GIc were purified and obtained from Arabidopsis plant extracts (Nakabayashi et al., 2008). Example 4: Tissue specificity of saiginol accumulation Plant material

Arabidopsis C24 plants were cultured on agar plates in a growth chamber under normal long day light conditions (16 h day, 140-160 μηηοΙ photons m ^"2 s ^"1 , 20°C; 8 h night, 16°C) for 14 days and transferred to soil. Tissue materials were harvested and immediately frozen in liquid nitrogen, and stored at -80°C until further use. Relative amount was compared and evaluated using detected peak area normalized by fresh weight material. The inventive saiginols were predominantly present in floral tissues but also to a lesser extent in the stem, silique and the cauline leaf but are essentially, or maybe even totally, absent in rosette leaves, senescent leaves and the root (Fig. 4) suggesting that they accumulate in the tissues which are highly exposed to light irradiation and most important with respect to facility. Further detailed spatial analysis of flavonol profiles revealed that the inventive saiginols predominantly accumulate in the floral petal and to a lesser extent in the stamen and pistil, but are probably absent in pollen.

Example 5: Metabolite Quantitative Trait Loci (mQTL) analysis for saiginols

The inventors aimed to narrow down the genomic basis for the capability of accession C24 to produce the inventive saiginols, compared to the accession ColO that is not capable to produce the inventive saiginols. Therefore, near Isogenic Lines (NILs) were obtained from the saiginol-producing accession C24 and the non-producing accession ColO by reciprocal introgression as described previously (Torjek et al., 2008, J. Hered. 99, 396- 406).

In brief: In order to create a desired NIL, a plant having the phenotype of interest (here accession C24 capable of saiginol production, also called "donor") is crossed with an accession not capable of saiginol production (here ColO, also called "background"). The result is a F1 generation having a genome that consists of 50% of the genome of C24 and 50% of the genome of ColO. During gametogenesis of this F1 generation, crossing-over occurs leading to ColO-derived chromosomes with substitutions from C24 and C24-derived chromosomes having substitutions from ColO. Several generations of backcrossing with the ColO parent line (also called background) leads to daughter generations with a more and more reduced amount of genomic segments from C24 within the ColO background. This process is also called reciprocal introgression. Here, Arabidopsis lines derived from ColO containing genomic fragments of C24 were named "N lines". Vice versa, it is also possible to backcross the F1 generation with the C24 line leading to daughter generations with a more and more reduced amount of genomic segments from ColO within the C24 background. In this case, the accession C24 would be the background line, and the accession ColO the donor. Here, Arabidopsis lines derived from C24 containing genomic fragments of ColO were named "M lines".

In total, 45 M lines (C24 background) and 69 N lines (ColO background) were used for saiginol targeted mQTL analysis. The presence or absence of the inventive saiginols in the NILs was assessed by LC-MS (compare Example 1 and 2).

Intriguingly, the inventors found a loss-of-function line in the NILs having a C24 background, i.e. a M line that is not capable of saiginol production (see NIL M16, Fig. 5 and Fig. 6 b), wherein the substituted genome segment (provided by ColO) mapped to chromosome 2 (At2g22230-At2g31610) as shown in Fig. 4.

Interestingly a genomic substitution (provided by C24) in the same chromosomal segment led to a gain-of-function line within the NILs having a ColO background, i.e. a N line that is now capable of saiginol production (see NIL N09, Fig. 5 and Fig. 6 a). Thus it can be concluded that a gene of accession C24 residing on chromosome 2 (At2g22230- At2g31610) is responsible to the production of the inventive saiginols.

O. Torjek et al. (2008): Construction and analysis of 2 reciprocal Arabidopsis introgression line populations. J. Hered. 99, 396-406.

Example 6: Transcript profiling via microarrav analysis

The inventors further performed transcript profiling experiments in several accessions producing the inventive saiginols (among them C24) and in several accession not producing the inventive saiginols (among them ColO) in order to compare the gene expression between producers and non-producers in the region of chromosome 2 assigned to be relevant for saiginol production (see Example 2) to narrow down the genetic basics for saiginol production Transcriptome analysis was carried out using ATH1 microarrays as described previously. The experiment was performed with five producing accessions (C24, Cvi, Da, Rsch and RLD) and five non-producing accessions (ColO, La-er, Ws, Sap and Stw).

In brief, the total RNA was extracted from frozen plant materials (see above) of each accession used by using the RNeasy Plant Mini Kit (Qiagen) according to the manufacturer's instructions.

Subsequently, labelled target cRNA was prepared according to the manufacturer's instructions of the Arabidopsis Genome ATH1 DNA array (Affymetrix, Santa Clara, CA, USA). For this, first double-stranded cDNA was prepared from 40 g of total RNA by use of the Superscript Choice System (invitrogen), again according to the manufacturer's instructions. Said cDNA was then transcribed in vitro using BioArray High Yield RNA Transcript Kit (Enzo, New York, NY, USA) into labelled cRNA. Following purification and fragmentation, the labeled cRNA was hybridized to Arabidopsis Genome ATH1 GeneChip array in a Hybridization Oven model 640 (Affymetrix). Washing and staining of the GeneChips were carried out using GeneChip Fluidics Station model 400. Scanning of the GeneChips was carried out with gene Array Scanner (Agilent Technologies). Duplicate hybridisations were carried out for ColO and C24, while for all other accessions a single hybridization was performed.

As a result, the expression levels were generally very similar between the genotypes although a number of genes were significantly different between producing and non- producing accessions including several transposable elements. However, only two genes were dramatically altered between ColO and C24, whereas the remaining 827 genes, which were localized to the chromosomal segment of interest exhibited regular expression (see Fig. 7). The genes altered in their expression between ColO and C24 were renamed as putative flavonol Qhenylacyltransferases 1 and 2 (FPT1 and FPT2). The gene transcripts for FPT1 and FPT2 were more than 17.7fold and 6.7fold higher in accession C24 than in accession ColO.

Example 7: Cloning of full-length cDNA of FPT1 and FPT2 genes from ColO and C24 Arabidopsis

The C24 and ColO alleles of both genes (FPT1 and FPT2) were cloned and the transcript production was evaluated. For this, first the total RNA was extracted from frozen plant materials (see above) of both accessions ColO and C24 by using the RNeasy Plant Mini Kit (Qiagen©) according to the manufacturer's instructions.

The mRNA contained in the total RNA that was used as template, was transcribed into oligo(dT)-primed cDNA by using using Superscript III Reverse Transcriptase (Invitrogen©, Germany) according to the manufacturer's protocol. After digestion of the remaining RNA by an RNase H treatment, 1/10 of the cDNA reaction volume was used for PCR with gene- specific primers for FPT2 (SEQ-ID No. 5 & 6) or with gene-specific primers for FPT1 (SEQ-ID No. 19 & 20).

The obtained PCR products were purified, cloned into a bacterial vector, transformed into E. coli, expressed in E. coli and purified. A cDNA segment was subcloned into GATEWAY pENTR Dual Selection vector (Invitrogen©, Germany). Then, 50ng of the resulting plasmid expressing are mixed with 50μΙ of Escherichia coli One Shot® TOP10 Chemically Competent E. coli cells (Invitrogen©, Germany) and incubated on ice for 10min. The mix is subsequently heated to 42°C for 45 seconds followed by 5min incubation in ice. 1 ml of Lysogeny Broth (LB) medium is added before incubation at 37°C for 1 h. Bacteria transformed in such a way are then plated on LB-agar plates containing 50 g/ml of kanamycin for selection of positive clones. A positive clone was grown in LB medium over night. Afterwards the cells were pelleted and the expressed plasmids containing the cloned cDNA fragment were isolated using the kit NucleoSpin Plasmid (Macherey-Nagel, Germany). Finally, the fragments were sequenced.

As a result, FPT1 transcript was invariant between ColO and C24 (see SEQ-ID No. 3). However, the FPT2-ColO allele encoded four different transcripts: FPT2-1 (ColO) (737 bp in length, SEQ-ID No. 7), FPT2-2(ColO) (651 bp in length, SEQ-ID No. 8), FPT2-3(ColO) (631 bp in length, SEQ-ID No. 9) and FPT2-4(ColO) (560 bp in length, SEQ-ID No. 10). In contrast, FPT2-C24 encoded a single but considerably longer transcript (1305 bp, see SEQ-ID No. 1 ).

Example 8: Genomic sequence of FPT2 gene in Arabidopsis accessions

Primers used for amplification and sequencing of FPT2 are described below. Four primer sets for both producing/non-producing accessions, two primer sets for only producing accessions were used. DNA fragments amplified by sets of primers were sequenced by ABI PRISM® 3700 Genetic Analyzer (Applied Biosystems, USA)

Genomic sequence analyses of FPT2 revealed a large gene deletion (-2279 bp) in ColO, spanning the region corresponding to the second to tenth exons of FPT2-C24. This analysis indicated that the four transcripts of FPT2-ColO described above (see Example 6) are alternative splicing variants.

Example 9: Overexpression of FPT2 and FPT1 in ColO plants

In order to experimentally test the function of FPT1 and FPT2 in vivo, the inventors performed complementation assays in ColO accessions by introducing an expression vector containing one of the transcripts described in Example 6 and over-expressing these transcripts.

FPT1 (Col0) and FPT1 (C24), indicate FPT1 cloned from ColO and C24, respectively.

FPT2-1 (ColO), FPT2-2(ColO) and FPT2(C24) indicate FPT2 cloned from ColO (FPT2-1 is shortest transcript and FPT2-2 is longest transcript) and C24, respectively.

A cDNA segment was subcloned into GATEWAY pK2GW7 vector (Invitrogen©, Germany). Then, obtained plasmids were transformed to Agrobacterium tumefaciens strain GV3101 pmp 90. Positive clones, which were checked by kanamycin selection, were suspended in 20 μΙ of sterile water and plated out on YEB petridishes with gentamycin (25 mg/l), rifampicin (100 mg/l) and the bacterial resistance marker kanamycin (50 mg/l). After incubation for 48 hours at 28°C, the solution was mixed with transformation medium (5% sucrose medium) for flower dipping transformation. T1 seedling plants have been selected by MS-agar plates (Murashige-Skoog-Medium agar plates) containing 50 g/ml of kanamycin for selection of positive clones. These antibiotics selection were repeated for T2 and T3 generation seedling to obtain T3 homozygote transgenic plants. In order to determine the saiginol production in the transformed plants, flower and leaf materials of Col-0, NIL N09, tt4 mutant and transgenic lines (FPT2(C24)-OX, FPT2- 1 (Col0)-OX, FPT1 (C24)-OX) were harvested for saiginol profile by chromatographic methods as described previously. As a result, overexpression of FPT1 , irrespective if derived from C24 (FPT1 (C24)) or ColO (FPT1 (Col0)), did not lead to a production of saiginols in ColO plants (see Fig. 8) The same result was obtained for the overexpression of the shortest transcript (FPT2-1 (ColO)), or the longest transcript (FPT2-2(ColO)) of FPT2 derived from ColO (Fig. 8). Only the overexpression of FPT2 derived from C24 (FPT2(C24)) led to the production of saiginols in the otherwise non-producing accession ColO (Fig. 8).

Example 10: UV-B irradiation experiment

In order to test the effect of saiginol production with respect to UV-B irradiation in vivo, the following plants were tested in this experiment:

(1 ) ColO as a representative for a non-producing accession

(2) NIL N09 - the gain-of-function line (C24 donor, ColO background, Example 5)

(3) FPT2(C24)-OX - transgenic ColO plant overexpressing FPT2(C24) (Example 8)

(4) FPT2-1 (ColO)-OX - transgenic ColO plant overexpressing FPT2-1 (ColO) (Example 8) (5) FPT1 (C24)-OX - transgenic ColO plant overexpressing FTP1 (C24) (Example 8)

(6) tt4 - a mutant Arabidopsis line lacking flavonoids Plants were cultured on agar plate in a growth chamber under normal long day light conditions (16 h day, 140-160 μηηοΙ photons m ^"2 s ^"1 , 20°C; 8 h night, 16°C) for 14 days and transferred to soil.

One week after transferring the plants to the soil and before plant bolting, plants were started to be irradiated with UV-B light in addition to the normal light regimen. UV-B light was irradiated 2h per day during midday time (stated 5-7h after starting normal light). The intensities of UV-B light tested in two trials were 1000 mW/m ² and 2000 mW/m ² .

After bolting of the plants, the seeds were collected and both the seed yield and the seed germination rate were determined using 2 weeks old seedling.

The seed yield was calculated as ratio between the seed yield (in mg) of an UV-B radiated plant and the seed yield (in mg) of a control plant without additional UV-B irradiation.

Not surprisingly, the flavonoid less tt4 mutant line was especially prone to UV-B irradiation and showed the lowest seed yield and seed germination rate (Fig. 9b, c). The seed yield of the non-producing accession ColO was reduced in comparison to not UV-B irradiated ColO plants, especially with higher UV-B intensity (Fig. 9b). The ColO lines overexpressing FPT1 (C24) or FPT2-1 (ColO) were indistinguishable from regular ColO (Fig. 9b). However, the seed yield in the gain-of-function line NIL N09 and the ColO line overexpressing FPT2(C24) was considerably higher compared to normal ColO plants (Fig. 9b). Moreover, the germination rate of the seeds was also doubled in NIL N09 and FPT2(C24)-OX compared to ColO (Fig. 9c), revealing that the production of germinable seeds was almost five times higher in the gain-of-function (NIL N09) line and the C24- FPT2 overexpressing line than in lines that do not produce the inventive saiginols.

In addition, the vitality of the NIL N09 gain-of-function line and the FPT2-C24 overexpressing line was reflected in the number of leaves and inflorescences these lines produced compared to ColO and tt4 (Fig. 9a). Moreover, NIL N09 and FPT2-C24 overexpressing line was considerably larger than tt4.

For silique production experiment (result shown in figure 10), inflorescences from the primary bolts of 5-week-old Arabidopsis plants were used. The detached inflorescences were irradiated with UV-B light (1 W m2) for 2 h per day during midday time (5-7 h after the onset of normal light) for 14 days by placing their cut ends in wells of a 96-well microtiter plate containing water.

Example 11 : Sunscreen formulation

A suitable sunscreen composition contains the following components with listed percentages based on total weight of the composition:

Active ingredients:

- titanium oxide (20.0%),

- saiginol A (7.5%),

Inactive ingredients:

- ethanol (59.9%),

- hyaluronic acid (2.5%),

- grape seed oil (5.0%),

- sesame oil (2.5%),

- sunflower seed oil (2.5%), and

- tetrahexyldecyl ascorbate (0.1 %).

Example 12: Transformation of tobacco and tomato plants

A cDNA segment was subcloned into GATEWAY pK2GW7 vector (Invitrogen©, Germany). Then, obtained plasmids were transformed to Agrobacterium tumefaciens strain GV2260 pmp 90. Positive clones, which were checked by spectinomycin selection, were suspended in 20 μΙ of sterile water and plated out on YEB petridishes with gentamycin (25 mg/l), rifampicin (100 mg/l) and the bacterial resistance marker spectinomycin (50 mg/l). After incubation for 24 hours at 28°C, the solution was mixed with transformation medium (5% sucrose medium) for transformation. Tobacco or tomato leaf disc (1x1 cm) which was injured by metal blade was used for injection. After dipping into agrobacterium solution, leaf discs were incubated on MS-agar plate for 48 h under darkness. After this treatment, leaf disc has been transferred to MS-agar plate (Murashige-Skoog-Medium agar plates) containing 50 g/ml of kanamycin for selection of positive clones and 6- Benzylaminopurine (BAP) and indole-3-acetic acid (IAA) for induction of callus root.

Example 13: Secondary metabolite profiling of transgenic tobacco plants by LC-MS Application of liquid chromatographyDmass spectrometry (LC-MS) using the method described in Example 2 resulted in the detection of a total of 3 peaks annotated as putative phenyacylated flavonol-glycosides in tobacco. All data were processed using Xcalibur 2.1 software (Thermo Fisher Scientific, Waltham, USA). This result indicates that FPT2 gene can be employed to metabolic engineering in other plant species. Since function of FPT2 has a broader function with lower substrate specificity, it can be used to transform other plant species than Arabidopsis and improve their UV-B resistance. SEQUENCE LIST

SEQ-ID No. 1: cDNA of C24 FPT2

ATGAGAACTTTTTCACCCAAGTTGCTGCTTCTTCTTTTACTTGTTTTAAGACATCATGCT GAATCTGGCTCTA TCGTCAAGTTTCTTCCCGGTTTTGAAGGCCCTCTTCCTTTCGAACTTGAAACCGGGTACA TCGGTATTGGTGA AGAAGAAGAACTGCAATCGTTTTACTATTTCATTAAGTCTGAGAAGAATCCAAAAGAAGA TCCTCTTCTTCTT TGGATATCTGGAGGACCTGGTTGCTCTTCTATTTCTGCTCTTCTTTTTGAGAATGGACCT GTGGCTCTAAAGT TCGAGGTTTACAATGAAACTCTCCCTTATTTGGTCTCTACTACATATTCATGGACCAAGA TGACGAACGTATT ATTCTTGGATCAGCCTATTGGAGTTGGCTTCTCCTACAAAAGAACTCCAAATCTTGATAA ATCGAGTGACACA ATAGAAGTATTGCGGATATACGAATTTCTTCAGAAGTGGCTAGGTGAACATCCTGAGTTT TTCTCCAACACTT TTTACGTAGGAGGAGATTCTTATTCCGGTAAGATTGTTCCAGCTATCGTTGATAAAATCT CACAAGAAAATTA TTTGTGCTGCAAACCTCCAATAAATCTTCAGGGTTATGTTCTCGGAAACCCAATAACAAA TTTGGAATCTGAT TCTAACTATCGTATTCCATATGCTCATGGAATGGCATTAATTTCTGATGAGCTCTACGAA TCCCTGAAGAGAA ACTGCAAAGGAAGATATAAAACCGTGGATCCATCTAACAAAAAATGTTTGAAACTTGTTG AAAAATACAATAA GTGTTCTGATAAAATATTTAGAGAACTAATATTATTACCACAGTGTGATGAAAGATCTCC ACTCTGCTGGGGC TACCACAGTACACTAGCTAAATATTGGGCCAATGACGAGAGGGTACGTGAAGCTCTTCAA ATAAGAAAGGGAA GTATAGGAAAATGGATACGATGTAATACGAATATACATTACGGTGACGACATTATTAGCA GCATACCATATCA TATGAATAACAGCATCAACGGATACCGATCTCTCATTTACAGTGGTGATCACGATATGGA GGTACCTTTCCTT GCAACTGAAGCTTGGATAAGATCTCTCAATTATCCTATTATTGATGATTGGAGGCCTTGG ATAATAAACAATC AGATTTCAGGATACACGATGACCTATGCCAATAAGATGACATATGCTACTATCAAGGGAG GTGGACACACTGC AGAGTATAAACCAGCGGAGAGCTTTATCATGTTCCAACGATGGATCAGTGGCCAGTCTCT GTAA

SEQ-ID No. 2: amino acid sequence coded by SEQ-ID No. 1

MRTFSPKLLLLLLLVLRHHAESGSIVKFLPGFEGPLPFELETGYIGIGEEEELQSFYYFI KSEKNPKEDPLLL WISGGPGCSSI SALLFENGPVALKFEVYNETLPYLVSTTYSWTKMTNVLFLDQPIGVGFSYKRTPNLDKSS DT IEVLRIYEFLQKWLGEHPEFFSNTFYVGGDSYSGKIVPAIVDKI SQENYLCCKPPI LQGYVLGNPITNLESD SNYRI PYAHGMALISDELYESLKRNCKGRYKTVDPSNKKCLKLVEKYNKCSDKI FRELILLPQCDERSPLCWG YHSTLAKYWANDERVREALQIRKGSIGKWIRCNTNIHYGDDI I SSI PYHMNNSINGYRSLIYSGDHDMEVPFL ATEAWIRSLNYPI IDDWRPWI INNQI SGYTMTYANKMTYATIKGGGHTAEYKPAESFIMFQRWI SGQSL

SEQ-ID No. 3: cDNA of C24 FPTl

ATGAAATCAACACTAAAATTGCTGCTTCTGCTTCTGTTTATGTTAAACCATCATGTTGAT TCTGGCTCTATCG TCAAGTTTCTTCCCGGCTTTGAAGGCCCTCTTCCTTTCGAACTCGAAACCGGGTACATTG GTATTGGTGAGGA AGAGGAAGTACAGTTGTTCTACTACTTTATAAAGTCTGAGAGAAATCCAAAAGAAGACCC TCTTCTTCTCTGG TTAAGTGGAGGACCTGGATGTTCATCTATCACTGGCCTTCTTTTCGAGAATGGACCTTTG GCTTTGAAGTCCG AGGTTTACAATGGAAGTGTCCCTTCTTTGGTCTCTACTACATATTCGTGGACAAAGACGG CGAACATAATATT CTTGGATCAGCCTATTGGAGCTGGCTTCTCCTACTCAAGAATCCCACTTATTGATACGCC TAGTGACACAGGC GAAGTTAAGAATATCCATGAGTTTCTCCAAAAGTGGTTAAGCAAGCATCCACAGTTTTCT TCCAATCCTTTCT ATGCTAGCGGAGATTCTTATTCCGGTATGATTGTTCCAGCCCTCGTTCAAGAAATTTCGA AAGGAAATTATAT ATGTTGCAAACCTCCTATAAATCTACAGGGCTATATACTCGGGAACCCAATAACATATTT TGAAGTCGACCAA AACTATCGCATTCCATTTTCTCATGGAATGGCACTTATTTCAGATGAACTATACGAGTCA ATTAGGAGAGACT GCAAAGGAAATTATTTCAACGTGGATCCACGTAACACAAAATGTTTGAAACTTGTTGAAG AATACCATAAGTG TACCGACGAACTAAATGAATTCAATATATTATCACCAGATTGCGACACGACATCTCCTGA TTGCTTTGTATAT CCATATTATCTCCTTGGCTACTGGATCAACGACGAGAGCGTTCGCGATGCTCTTCATGTT AATAAGAGCAGTA TTGGAAAATGGGAGCGATGTACTTATCAAAATAGAATCCCATACAACAAAGACATCAATA ACAGCATACCATA CCATATGAATAACAGTATTAGTGGCTACCGATCTCTCATCTACAGTGGTGATCATGATTT GGTGGTTCCTTTC CTTGCAACTCAAGCCTGGATAAAATCTCTAAATTACTCCATCATTCATGAATGGAGACCT TGGATGATTAAAG ATCAAATCGCTGGGTATATAATATATTTTTGTGTTATACACGAGAACTTATTCCAATAA

SEQ-ID No. 4: amino acid sequence coded by SEQ-ID No. 3

MKSTLKLLLLLLFMLNHHVDSGSIVKFLPGFEGPLPFELETGYIGIGEEEEVQLFYYFIK SERNPKEDPLLLW LSGGPGCSSITGLLFENGPLALKSEVYNGSVPSLVSTTYSWTKTANI I FLDQPIGAGFSYSRIPLI DTPSDTG EVKNIHEFLQKWLSKHPQFSSNPFYASGDSYSGMIVPALVQEI SKGNYICCKPPINLQGYILGNPITYFEVDQ NYRIPFSHGMALI SDELYESIRRDCKGNYFNVDPRNTKCLKLVEEYHKCTDELNEFNILSPDCDTTSPDCFVY PYYLLGYWINDESVRDALHVNKSSIGKWERCTYQNRIPYNKDINNSIPYHMNNSISGYRS LIYSGDHDLVVPF LATQAWIKSLNYSI IHEWRPWMIKDQIAGYI IYFCVIHENLFQ

SEQ-ID No. 5: forward primer for amplification of FPT2 cDNA (At2g22960)

ATGAGAACTTTTTCACCCAAGTT

SEQ-ID No. 6: reverse primer for amplification of FPT2 cDNA (At2g22960)

TCATCATCTCTTATTATTACAGA

SEQ-ID No. 7: cDNA of ColO FPT2-1

ATGAGAACTTTTTCACCCAAGTTGCTGCTTCTTCTTTTACTTGTTTTAAGACATCATGCT GAATCTGGCTCTA TCGTCAAGTTTCTTCCCGGTTTTGAAGGCCCTCTTCCTTTCGAACTTGAAACCGGGGCTA CCACAGTACACTA GCTAAATATTGGGCCAATGACGAGAGGGTACGTGAAGCTCTTCAAATAAGAAAGGGAAGT ATAGGAAAATGGA TACGATGTAATTCGAATATACATTACGATGACGACATTATTAGCAGCATACCATATCATA TGAATAACAGCAT CAACGGATACCGATCTCTTATTTACAGGTTTGATTAAAATTACATTTTAAAGAATCGATC TAGTTCTCTATAC AAACTATGCGGTTTTTGGTAAACAGTGGTGATCACGATATGGAGGTACCTTTCCTTGCAA CTGAAGCTTGGAT AAGATCTCTTAATTATCCTATTATTGATGATTGGAGGCCTTGGATAATAAACAATCAGAT TGCTGGGTGAATA ATTTTTACAATTTTTTTTTTTGCTTCTTTATTCATTAACTCAGTCGAATCATGAGCTCAT TAGTCATTTTGAA ATCCCTCTTACGATTTTCTTGGACAGATACACGATGACCTATGCCAATAAGATGACATAT GCTACTATCAAGG GAGGTGGACACACTGCAGAGTATAAACCAGCGGAGAGCTTTATCATGTTCCAACGATGGA TCAGTGGCCAGCC TCTGTAA

SEQ-ID No. 8: cDNA of ColO FPT2-2

ATGAGAACTTTTTCACCCAAGTTGCTGCTTCTTCTTTTACTTGTTTTAAGACATCATGCT GAATCTGGCTCTA TCGTCAAGTTTCTTCCCGGTTTTGAAGGCCCTCTTCCTTTCGAACTTGAAACCGGGGCTA CCACAGTACACTA GCTAAATATTGGGCCAATGACGAGAGGGTACGTGAAGCTCTTCAAATAAGAAAGGGAAGT ATAGGAAAATGGA TACGATGTAATTCGAATATACATTACGATGACGACATTATTAGCAGCATACCATATCATA TGAATAACAGCAT CAACGGATACCGATCTCTTATTTACAGTGGTGATCACGATATGGAGGTACCTTTCCTTGC AACTGAAGCTTGG ATAAGATCTCTTAATTATCCTATTATTGATGATTGGAGGCCTTGGATAATAAACAATCAG ATTGCTGGATACA CGATGACCTATGCCAATAAGATGACATATGCTACTATCAAGGCAAGTTTTTTTTTTTTGT TCTTACTCTTAAT GTTTTTTATTACACAAATCATTTTGTTTCTCAGTAACTGTTTTGAGGGGTTTACTATAGG GAGGTGGACACAC TGCAGAGTATAAACCAGCGGAGAGCTTTATCATGTTCCAACGATGGATCAGTGGCCAGCC TCTGTAA

SEQ-ID No. 9: cDNA of ColO FPT2-3

ATGAGAACTTTTTCACCCAAGTTGCTGCTTCTTCTTTTACTTGTTTTAAGACATCATGCT GAATCTGGCTCTA TCGTCAAGTTTCTTCCCGGTTTTGAAGGCCCTCTTCCTTTCGAACTTGAAACCGGGGCTA CCACAGTACACTA GCTAAATATTGGGCCAATGACGAGAGGGTACGTGAAGCTCTTCAAATAAGAAAGGGAAGT ATAGGAAAATGGA TACGATGTAATTCGAATATACATTACGATGACGACATTATTAGCAGCATACCATATCATA TGAATAACAGCAT CAACGGATACCGATCTCTTATTTACAGGTTTGATTAAAATTACATTTTAAAGAATCGATC TAGTTCTCTATAC AAACTATGCGGTTTTTGGTAAACAGTGGTGATCACGATATGGAGGTACCTTTCCTTGCAA CTGAAGCTTGGAT AAGATCTCTTAATTATCCTATTATTGATGATTGGAGGCCTTGGATAATAAACAATCAGAT TGCTGGATACACG ATGACCTATGCCAATAAGATGACATATGCTACTATCAAGGGAGGTGGACACACTGCAGAG TATAAACCAGCGG AGAGCTTTATCATGTTCCAACGATGGATCAGTGGCCAGCCTCTGTAA

SEQ-ID No. 10: cDNA of ColO FPT2-4

ATGAGAACTTTTTCACCCAAGTTGCTGCTTCTTCTTTTACTTGTTTTAAGACATCATGCT GAATCTGGCTCTA TCGTCAAGTTTCTTCCCGGTTTTGAAGGCCCTCTTCCTTTCGAACTTGAAACCGGGGCTA CCACAGTACACTA GCTAAATATTGGGCCAATGACGAGAGGGTACGTGAAGCTCTTCAAATAAGAAAGGGAAGT ATAGGAAAATGGA TACGATGTAATTCGAATATACATTACGATGACGACATTATTAGCAGCATACCATATCATA TGAATAACAGCAT CAACGGATACCGATCTCTTATTTACAGTGGTGATCACGATATGGAGGTACCTTTCCTTGC AACTGAAGCTTGG ATAAGATCTCTTAATTATCCTATTATTGATGATTGGAGGCCTTGGATAATAAACAATCAG ATTGCTGGATACA CGATGACCTATGCCAATAAGATGACATATGCTACTATCAAGGGAGGTGGACACACTGCAG AGTATAAACCAGC GGAGAGCTTTATCATGTTCCAACGATGGATCAGTGGCCAGCCTCTGTAA

SEQ-ID No. 11: forward primer of set 1 for amplification of FPT2 genome sequence

TGGACTAGTACGAGAATTGCAAAG

SEQ-ID No. 12: reverse primer of set 1 for amplification of FPT2 genome sequence

ACCGGGAAGAAACTTGACGA

SEQ-ID No. 13: forward primer of set 2 for amplification of FPT2 genome sequence

CGTGGAGCGACCAGTGAT

SEQ-ID No. 14: reverse primer of set 2 for amplification of FPT2 genome sequence

CTTCACGTACCCTCTCGTCA SEQ-ID No. 15: forward primer of set 3 for amplification of FPT2 genome sequence

CTTATGCAGGGCTACCACAGT

SEQ-ID No. 16: reverse primer of set 3 for amplification of FPT2 genome sequence

ACCACTGTTTACCAAAAACCGC SEQ-ID No. 17: forward primer of set 4 for amplification of FPT2 genome sequence

ATATCATATGAATAACAGCATC

SEQ-ID No. 18: reverse primer of set 4 for amplification of FPT2 genome sequence

GCATAGGGCATCATCATCTC

SEQ-ID No. 19: forward primer for amplification of FPTl cDNA

(At2g22920)

ATGAAATCAACACCAAAAT

SEQ-ID No. 20: reverse primer for amplification of FPTl cDNA

(At2g22920)

ATTTAGGTAAGGTCATTCAAGTA

SEQ-ID No. 21: amino acid (aa) sequence of a variant of BLOCK 1

GPGCSSxxxL

(x = random amino acid)

SEQ-ID No. 22: amino acid (aa) sequence of a variant of BLOCK 2

GDSYSG