RRC ID 36142
Author Zhu L, Mon H, Xu J, Lee JM, Kusakabe T.
Title CRISPR/Cas9-mediated knockout of factors in non-homologous end joining pathway enhances gene targeting in silkworm cells.
Journal Sci Rep
Abstract Gene targeting can be achieved by precise genetic modifications through homology-directed repair (HDR) after DNA breaks introduced by genome editing tools such as CRISPR/Cas9 system. The most common form of HDR is homologous recombination (HR). Binding to the DNA breaks by HR factors is thought to compete with non-homologous end joining (NHEJ), an alternative DNA repair pathway. Here, we knocked out the factors in NHEJ by CRISPR/Cas9 system in silkworm cells, so that increased the activities of HR up to 7-fold. Also efficient HR-mediated genome editing events occurred between the chromosomal BmTUDOR-SN gene and donor DNA sequences with an EGFP gene in the middle of two homologous arms for the target gene. Utilizing the NHEJ-deficient silkworm cells, we found that homologous arms as short as 100 bp in donor DNA could be designed to perform precise genome editing. These studies should greatly accelerate investigations into genome editing of silkworm.
Volume 5
Pages 18103
Published 2015-12-10
DOI 10.1038/srep18103
PII srep18103
PMID 26657947
PMC PMC4674802
MeSH Animals Animals, Genetically Modified Base Sequence Bombyx / cytology Bombyx / genetics* CRISPR-Cas Systems* Cells, Cultured DNA End-Joining Repair* Gene Knockout Techniques / methods* Gene Targeting / methods* Genetic Engineering / methods Genome, Insect / genetics Green Fluorescent Proteins / genetics Green Fluorescent Proteins / metabolism Homologous Recombination Insect Proteins / genetics Insect Proteins / metabolism Microscopy, Fluorescence Molecular Sequence Data Signal Transduction / genetics
IF 3.998
Times Cited 15