牦牛CART基因克隆、单核苷酸多态性检测及生物信息学分析

doi:10.11843/j.issn.0366-6964.2013.08.007

摘要/Abstract

摘要：

旨在克隆牦牛可卡因-苯丙胺调节转录肽基因（Cocaine- and amphetamine-regulated transcript，CART），获得基因全长序列并分析基因结构和相关遗传变异，检测单核苷酸多态性（SNP）位点在不同品种牦牛群体中的分布，通过生物信息学相关方法分析预测SNP位点对CART基因表达的影响，为牦牛CART基因功能研究奠定基础。以243头大通牦牛、208头天祝牦牛、208头甘南牦牛和53头帕里牦牛血样为材料，提取基因组DNA，克隆出牦牛CART基因全长序列；运用高分辨率熔解曲线分析技术（High-resolution melting, HRM）检测CART基因4个SNPs在4个牦牛品种中的分布，进行遗传统计分析，并应用生物信息学分析SNPs可能对基因功能产生的影响。本研究发现，牦牛CART基因4个SNPs，1个位于基因上游序列、1个位于内含子、2个位于3′-UTR。Χ²检验表明，除大通牦牛V-221位点和甘南牦牛V1816位点外，其他所有群体各个位点都处于哈代-温伯格平衡（P>0.05）。群体遗传学分析显示，V168和V1816位点属于中度多态，V-221和V1791属于低度多态，单倍型C属于优势单倍型。生物信息分析发现，V1791和V1816位点碱基变化导致RNA二级结构发生变化，可能会对CART基因的表达产生影响。

Abstract:

This study aimed to clone the complete length sequence of Cocaine- and Amphetamine-Regulated Transcript gene (CART), analyze its structure, and reveal the distribution of SNPs (single nucleotide polymorphisms) in different yak breeds. Possible impact of the SNPs on CART gene expression was also predicted by bioinformatics analysis. This study could provide some basic information for studying the function of yak CART gene in future. The blood samples of 243 Datong yak, 208 Tianzhu yak, 208 Gannan yak, and 53 Pali yak were used to isolate the genomic DNA. Then the full length of CART gene was cloned and sequenced. Genetic variations and the distribution of 4 SNPs in the 4 yak breeds were detected by the high-resolution melting system followed by genetic statistical analysis. In this study, 4 novel SNPs located in the yak CART gene were identified, and these SNPs were located in upstream, intron, and 3′-UTR of the CART gene. Χ² tests showed that besides the V-221 in Datong population and V1816 in Gannan population, all the other loci were in Hardy-Weinberg equilibrium (P＞0.05) in the 4 yak breeds. Through population genetics analysis, the results showed that V168 and V181 loci were at intermediate polymorphic status, but V-221 and V1791 loci were at low polymorphic status. The haplotype C was predominant. Bioinformatics analysis showed that different alleles of V1791 and V1816 might change the RNA secondary structure, which could influence CART gene’s expression. The nucleotide variation of V1791 and V1816 loci reduced RNA stability that might produce negative influence on CART gene expression.

中图分类号:

S813.3

刘建，刘文博，吴晓云，阎萍，梁春年. 牦牛CART基因克隆、单核苷酸多态性检测及生物信息学分析[J]. 畜牧兽医学报, 2013, 44(8): 1219-1228.

LIU Jian, LIU Wen-bo, WU Xiao-yun, YAN Ping, LIANG Chun-nian. Cloning, Single Nucleotide Polymorphism and Bioinformatics Analysis of CART Gene in Yak[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2013, 44(8): 1219-1228.

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