Title:
LINKED DNA PRODUCTION METHOD AND VECTOR COMBINATION FOR USE THEREIN
Document Type and Number:
WIPO Patent Application WO/2021/192596
Kind Code:
A1
Abstract:
This method for producing linked DNA in which DNA fragments are linked includes: (a1) a step for preparing a first vector, which contains a (1) 5'-R1-D(i)-R2-M1-R2'-D(ii)-R1'-3' structure containing restriction enzyme recognition sequences R1, R1', R2, and R2', selection marker genes M1 and M2, which are different from each other, and arbitrary linking DNA fragments D(i) through D(iv), and preparing a second vector, which contains a (2) 5'-R1'-D(iii)-R2-M2-R2'-D(iv)-R1'-3' structure; (b1) a step for treating the first vector with a first restriction enzyme and a second restriction enzyme, and subsequently obtaining a first vector fragment composed of a 5'-D(i)-R2-M1-R2'-D(ii)-3' structure; (c1) a step for treating the second vector with a third restriction enzyme and a fourth restriction enzyme, and subsequently obtaining a second vector fragment in which a 5'-R2-M2-R2'-3' structure has been removed; and (d1) a step for linking, via a ligation reaction, the first vector obtained in step b1 and the second vector obtained in step c1, and generating a third vector, which contains a (3) 5'-R1-D(i)1-R2-M1-R2'-D(ii)1-R1'-3' structure (here, D(i)1 denotes a DNA fragment containing a 5'-D(iii)-D(i)-3' structure, and D(ii)1 denotes a DNA fragment containing a 5'-D(ii)-D(iv)-3' structure).
Inventors:
YACHIE NOZOMU (JP)
MORI HIDETO (JP)
YAMAGUCHI NANAMI (JP)
MORI HIDETO (JP)
YAMAGUCHI NANAMI (JP)
Application Number:
PCT/JP2021/003060
Publication Date:
September 30, 2021
Filing Date:
January 28, 2021
Export Citation:
Assignee:
UNIV TOKYO (JP)
International Classes:
C12N15/64; C12N1/21
Domestic Patent References:
| WO2018203056A1 | 2018-11-08 |
Foreign References:
| JP2012509084A | 2012-04-19 |
Other References:
STORCH, M. ET AL.: "BASIC: A NEW BIOPART ASSEMBLY STANDARD FOR IDEMPOTENT CLONING PROVIDES ACCURATE, SINGLE-TIER DNA ASSEMBLY FOR SYNTHETIC BIOLOGY", ACS SYNTHETIC BIOLOGY, vol. 4, 17 July 2015 (2015-07-17), pages 781 - 787, XP055343133, DOI: 10.1021/sb500356d
Attorney, Agent or Firm:
CENTCREST IP ATTORNEYS (JP)
Download PDF: