高通量低成本检测牛MSTN基因突变的DAVID技术构建  

Construction of DAVID Technology for High-throughput and Low-cost Detection of Bovine (Bos taurus) MSTN Gene Mutations

作　　者：裴栋超 海超 赵春生 张立果 吴迪 王德政 张鑫怡 杨磊 李光鹏 PEI Dong-Chao;HAI Chao;ZHAO Chun-Sheng;ZHANG Li-Guo;WU Di;WANG De-Zheng;ZHANG Xin-Yi;YANG Lei;LI Guang-Peng(State Key Laboratory of Reproductive Regulation&Breeding of Grassland Livestock,Inner Mongolia University,Hohhot 010070,China)

机构地区：[1]内蒙古大学省部共建草原家畜生殖调控与繁育国家重点实验室,呼和浩特010070

出　　处：《农业生物技术学报》2025年第1期208-219,共12页Journal of Agricultural Biotechnology

基　　金：科技创新2030-重大项目(2023ZD0404803);国家自然科学基金(32360837;32341052);内蒙古科技领军团队(2022LJRC0006);农业农村部农业重大科技项目(NK202213020303);内蒙古揭榜挂帅项目(2022JBGS0025);内蒙古科技重大专项(2021ZD0009;2021ZD0008;2022ZD0008);内蒙古科技计划(2023KJHZ0028);内蒙古青年科技英才项目(NJYT23138);中央引导地方科技发展资金(2022ZY0212);内蒙古自然科学基金(2023MS03004);呼和浩特市高校院所协同创新(XTCX2023-06);内蒙古大学教育部工程中心课题(JYBGCSYS2022);新疆维吾尔自治区重大科技专项(2023A02011-1)。

摘　　要：肌肉生长抑制素(myostatin, MSTN)是骨骼肌生长发育的负调控因子,MSTN基因突变可促进动物肌肉发育。目前Sanger测序是检测MSTN基因突变的经典方法,但此方法存在检测通量低、可视化程度低和检测费用高等缺点。本研究利用二代测序技术,通过设计的特异性Barcode序列,能够一次性检测96头牛(Bos taurus)的MSTN基因突变情况。此外,根据测序Fastq结果文件中的Barcode序列,利用异步线程和并发算法,编写了相关的数据分析DAVID(detect and visualize insertion-deletion)软件。利用DAVID软件将96个混合样本的测序数据逐一拆分,并与扩增子序列(amplicon sequence)比对,最终获得MSTN基因突变的图形和统计信息。利用DAVID技术对96头蒙古牛进行检测,结果发现1头牛的MSTN基因11 bp杂合缺失,剩余95头均为野生型,并且通过Sanger测序对11 bp杂合缺失进行了验证。综上,本研究构建了一个检测牛MSTN基因突变的高通量技术体系,为肉牛遗传改良和育种提供技术支撑。Myostatin(MSTN)is a negative regulator of skeletal muscle growth and development,and mutations in the MSTN gene can promote muscle development in animals.Currently,Sanger sequencing technology is the classic method for detecting MSTN gene mutations,but this method has the disadvantages of low detection throughput,low visualisation of mutation information and high detection cost.In this study,MSTN gene mutations in 96 cattle(Bos taurus)could be detected at one time by using next-generation sequencing technology with a specific Barcode sequence designed.In addition,based on the Barcode sequences in the sequencing Fastq result files,the related data analysis DAVID(detect and visualize insertion-deletion)software was developed using asynchronous threading and concurrent algorithms.Using DAVID software,the sequencing data of 96 mixed samples were split one by one and compared with amplicon sequence,and finally the graphical and statistical information of MSTN gene mutation was obtained.Using DAVID technology,96 Mongolian cattle were examined,and found that 1 head of cattle was heterozygous for 11 bp deletion of the MSTN gene,and the remaining 95 head of cattle were wild-type,and the 11 bp heterozygous deletion was further verified by Sanger sequencing.This study constructed a high-throughput technical system for detecting bovine MSTN gene mutations,and provides technical support for genetic improvement and breeding of cattle.

关 键 词：肌肉生长抑制素(MSTN) 蒙古牛 基因突变 基因型检测 

分 类 号：S813.3[农业科学—畜牧学]