猪ZBED6基因有效突变位点的筛选与验证

发布时间：2018-03-04 13:04

本文选题：猪　切入点：ZBED基因　出处：《中国畜牧杂志》2017年08期 　论文类型：期刊论文

【摘要】：锌指蛋白ZBED6基因的突变可促进骨骼肌中胰岛素样生长因子2(IGF2)转录水平显著上调,从而促进细胞增殖和肌管形成。本研究利用CRISPR-Cas9技术对猪的ZBED6基因进行编辑,利用在线软件筛选出4个突变靶点,利用p Cas9/g RNA载体构建p Cas9/g RNA-ZBED6质粒,再构建4个用于验证突变效率的p TYNE-ZBED6质粒,同时转染HEK293细胞系,通过观察荧光信号的强弱判断靶序列突变效率的高低。结果表明:位于编码区第17~37位点的靶点1转染后荧光信号最强,切割效率最高,该位点是利用CRISPR-Cas9技术编辑ZBED6基因的有效位点。
[Abstract]:The mutation of zinc finger protein ZBED6 gene can significantly up-regulate the transcription level of insulin-like growth factor-2 IGF2 in skeletal muscle, thus promote cell proliferation and myotube formation. In this study, CRISPR-Cas9 technique was used to edit porcine ZBED6 gene. Four mutation targets were screened out by on-line software, then p Cas9/g RNA-ZBED6 plasmid was constructed by using p Cas9/g RNA vector, and four p TYNE-ZBED6 plasmids were constructed to verify the mutation efficiency. At the same time, they were transfected into HEK293 cell line. The mutation efficiency of target sequence was judged by observing the intensity of fluorescence signal. The results showed that target 1, located at site 1737 of coding region, had the strongest fluorescence signal and the highest cutting efficiency after transfection. This site is an effective site for editing ZBED6 gene by CRISPR-Cas9 technique.
【作者单位】：黑龙江省农业科学院畜牧研究所;农业部畜禽遗传资源与种质创新重点实验室;
【基金】：农业部畜禽遗传资源与种质创新重点实验室开放课题国家生猪产业技术体系(CARS-36)
【分类号】：Q78;S828

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