荷斯坦牛HH5遗传缺陷基因快速检测方法的建立及应用

HH5是新近发现的一种荷斯坦牛隐性遗传缺陷基因,其分子机理为9号染色体上138kb大片段缺失,造成二甲基腺苷转移酶1(TFB1M)基因丢失。TFB1M基因在核糖体小亚基合成中发挥重要功能,故缺陷纯合子胚胎在发育早期死亡,造成母牛流产。HH5基因的共同祖先为1957年出生的一头加拿大著名公牛,其因在世界范围内广泛扩散,奶业发达国家已将其列为常规检测的遗传位点。本研究首先通过Sanger测序分析HH5缺陷基因,发现除了已知的138kb大片段缺失外,还存在一个9bp插入片段(TGATTACAA);该基于此特征序列,设计等位基因特异性PCR分型方法,PCR产物经琼脂糖凝胶电泳即可快速分型。笔者所在团队对北京郊区某奶牛场372头荷斯坦母牛检测,发现HH5携带者12头,携带率为3.23%。本研究为我国荷斯坦牛HH5遗传缺陷基因筛查提供了一种准确便捷的分子检测方法。

Development and Application of An Efficient Molecular Assay for Genetic Defect HH5 in Holstein

HH5 is an autosomal recessively inherited disorder that was recently identified in Holstein cattle.The molecular mechanism of HH5 has been identified as a deletion of 138 kb region on chromosome 9,harboring dimethyl-adenosine transferase 1(TFB1M).Since TFB1M is essential for synthesis of the small ribosomal subunit of mitochondria,the embryo with homozygous defective alleles will die during early pregnancy.The HH5 defect was traced back to a famous Canadian Holstein bull born in 1957 and the mutant allele has spread worldwide.The current study aimed to develop an efficient molecular method for detecting HH5 genetic defect in Chinese Holstein cattle.We first identified an insertion of 9 bp in addition to the deletion of 138 kb in the HH5 defect allele by Sanger sequencing.Then an allele-specific PCR-based assay was established to discriminate the wild allele from the mutant allele.By using this new method,12 carriers were identified from 372 Holstein cows that were sampled in Beijing,corresponding to a carrier frequency of 3.23%.