RRC ID 54366
Author Nagasaki A, Kato Y, Meguro K, Yamagishi A, Nakamura C, Uyeda TQP.
Title A genome editing vector that enables easy selection and identification of knockout cells.
Journal Plasmid
Abstract The CRISPR/Cas9 system is a powerful genome editing tool for disrupting the expression of specific genes in a variety of cells. However, the genome editing procedure using currently available vectors is laborious, and there is room for improvement to obtain knockout cells more efficiently. Therefore, we constructed a novel vector for high efficiency genome editing, named pGedit, which contains EGFP-Bsr as a selection marker, expression units of Cas9, and sgRNA without a terminator sequence of the U6 promoter. EGFP-Bsr is a fusion protein of EGFP and blasticidin S deaminase, and enables rapid selection and monitoring of transformants, as well as confirmation that the vector has not been integrated into the genome. By using pGedit, we targeted human ACTB, ACTG1 and mouse Nes genes coding for β-actin, γ-actin and nestin, respectively. Knockout cell lines of each gene were easily and efficiently obtained in all three cases. In this report, we show that our novel vector, pGedit, significantly facilitates genome editing.
Volume 98
Pages 37-44
Published 2018-6-1
DOI 10.1016/j.plasmid.2018.08.005
PII S0147-619X(18)30085-4
PMID 30196057
MeSH Actins / antagonists & inhibitors* Actins / genetics Aminohydrolases / genetics Aminohydrolases / metabolism Animals Base Sequence CRISPR-Cas Systems* Gene Editing / methods* Gene Targeting Genetic Vectors* Green Fluorescent Proteins / genetics Green Fluorescent Proteins / metabolism Humans Mice Nestin / antagonists & inhibitors* Nestin / genetics Promoter Regions, Genetic Recombinant Fusion Proteins / metabolism* Sequence Homology
IF 2.805
Times Cited 4
DNA material pGedit (RDB16763)