山羊骨形态发生蛋白2基因外显子克隆及序列分析

Cloning and Sequence Analysis on Exons of Bone Morphogenetic Protein 2 Gene in Goats

设计4对引物,采用PCR-SSCP技术检测骨形态发生蛋白2（bone morphogenetic protein 2,BMP2）基因外显子2、3在济宁青山羊、安哥拉、波尔、内蒙古绒山羊4个品种共191个个体中的单核苷酸多态性,分析该基因对济宁青山羊高繁殖力的影响,并对济宁青山羊BMP2基因2个外显子进行克隆测序。结果表明,4对引物的扩增片段在4个山羊品种中均无多态性;山羊扩增片段核苷酸和氨基酸序列与人、黑猩猩、牛、犬、大鼠、小鼠6个物种的同源性分别为86.5%~91.3%和88.8%~99%;与其他物种相比,山羊BMP2扩增片段推导氨基酸序列存在3处特有突变,分别是位于第131、353、365位的脯氨酸、缬氨酸、亮氨酸变为苏氨酸（P131T）、天冬氨酸（V353D）、脯氨酸（L365P）。可见,哺乳动物BMP2基因外显子2、3序列保守性强,该区域可能不是影响山羊高繁殖力的功能结构域。

The bone raorphogenetic protein 2 （BMP2） gene was studied as a candidate gene for high prolificacy in some goat breeds （Capra hircus）. Single nucleotide polymorphisms （SNPs） of two exons （exons 2 and 3） of BMP2 gene were detected in high prolificacy breed （Jining G-rey goat）, moderate prolificacy breed （Boer goat） and low prolificacy breeds （Inner Mongolia Cashmere and Angora goats） by polymerase chain reaction-single strand conformation polymorphism （PCR-SSCP）. No polymorphism was detected in the amplified region in the four goat breeds tested. DNA fragments of exons 2 and 3 of Jining Grey goat BMP2 gene were cloned into pSRUE-T vector. The positive clones were further identified by PCR. The nucleotide sequences were detected and the amino acid sequences of these fragments were deduced. The comparison of the fragments of two exons of Jining Grey goat BMP2 gene with those of human, chimpanzee, cattle, dog, rat and mouse BMP2 gene showed that the homologies of the nucleotide sequences and amino acid sequences of the fragments among these seven species were from 86. 5% to91. 3% and from 88.8% to 99% respectively. Three special variations were found in the 131st, 353nd , 365th position of amino acid sequence, which was threonine, aspartic acid, proline in Jining Grey goats and proline, valine, leucine in other species respectively. The results indicated that the exons 2 and 3 of BMP2 gene were highly conserved among mammals, and may not be the functional domains that affect the high prolificacy in goats.