Construction and Identification of MSTN Knockout AAV-SaCas9
WANG Xinjian
FAN Shuangshuang
WENG Shaoting
YANG Guoyu
CHU Beibei
WANG Jiang
Abstract：To study the function of myostatin (MSTN),molecular biology method was applied to construct the recombinant vector of MSTN knockout AAV-SaCas9,the transfected 293T cells were extracted for the genome and identified by T7 endonuclease.Then the mutations were further confirmed by TA cloning and sequencing.The AAV-293 cells were co-transfected with AAV-SaCas9 and helper plasmids.Virus was isolated and purified after three days,and its titer was determined by real-time fluorescence quantitative PCR.Our results showed that the recombinant vector of MSTN knockout AAV-SaCas9 was successfully constructed.MSTN could be edited by sgRNA2 identified by T7 digestion and sequencing,and AAV-SaCas9 was successfully packaged into virus.Finally the titer was 2.73 × 1012 vg/mL,which was identified by fluorescence quantitative PCR.
Keywords：MSTN geneCRISPR-Cas9AAV vector293T cell
Publication Date：2018-01-01
Online Publishing Date：2026-09-12(First online date of this platform, not the publication date of the document)
Pages：4( 118-121 )