CRISPR/Cas技术可有效介导家鸡基因敲除

doi:10.11843/j.issn.0366-6964.2016.06.024

Abstract

Abstract:

This study aimed to establish a highly efficient and stable gene editing technique mediated by CRISPR/Cas to knock out the targeted gene，and explore its application in chicken preliminarily，and to provide the operation basis for the subsequent poultry genetic edition.We cloned the full-length of C2EIP (chr2，Expression in PGC) gene according to sequence in NCBI Database，and designed 3 gRNAs named gRNA1，gRNA2 and gRNA3 based on the location of APM in the sequence to construct the cas9/gRNA vector.SSA activity assay，T7E1 digestion method and sequencing of TA cloning were used to detect the knock-out efficiency of the gRNA after the cas9/gRNA vector transfected into DF-1.Results of SSA activity assay showed that only cas9/gRNA3 vector could knock out the gene effectively，fluorescence activity which transfected cas9/gRNA3 was twice higher than the control group；Result of T7E1 digestion showed that activity of cas9/gRNA3 was 27%，sequencing of TA cloning results showed there were 8 mutational samples in 30 samples，and the efficiency of gene-knockout was 26%.In this study we established the technology of gene knockout mediated by CRISPR/Cas in chicken，which can be used to konck out genes in DF-1 stably.

CLC Number:

S831.2

ZUO Qi-sheng，WANG Ying-jie，ZHAO Rui-feng，CHENG Shao-ze，WANG Yi-lin，JIN Kai，WANG Fei，JI Yan-qin，LU Zhen-yu，ZHANG Wen-hui，ZHANG Ya-ni，LI Bi-chun. CRISPR/Cas Techniques Can Knockout the Gene of Chicken Effectively[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2016, 47(6): 1266-1271.

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URL: https://www.xmsyxb.com/EN/10.11843/j.issn.0366-6964.2016.06.024

https://www.xmsyxb.com/EN/Y2016/V47/I6/1266

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CRISPR/Cas Techniques Can Knockout the Gene of Chicken Effectively

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