为了解促黄体素β基因多态性与济宁青山羊产羔数的关系,采用PCR-SSCP技术检测促黄体素β亚基(Luteinizing hormone betasubunit,LHβ)基因5'调控区和3 个外显子在高繁殖力(济宁青山羊)、中等繁殖力(波尔山羊)和低繁殖力山羊品种(辽宁绒山羊和安哥拉山羊)中的单核苷酸多态性,同时研究该基因对济宁青山羊高繁殖力的影响。结果表明:山羊与绵羊的LHβ基因核苷酸序列相似性为98%。6对引物中,仅引物P1和P5扩增片段存在多态性。对于P1扩增片段,在4个山羊品种中均检测到AA、AB和BB 3种基因型;测序分析发现BB与AA基因型相比在5'调控区存在2处突变202C→A和210C→T;济宁青山羊3种基因型之间产羔数差异均不显著(P〉0.05)。对于P5扩增片段,在济宁青山羊中检测到CC、CD和DD 3种基因型,辽宁绒山羊中检测到CC和CD 2种基因型,波尔山羊和安哥拉山羊中都只检测到CC基因型;测序分析发现DD与CC基因型相比在外显子2存在单碱基突变1124C→T;济宁青山羊CC、CD和DD基因型频率分别为0.38、0.44和0.18;DD和CD基因型济宁青山羊平均产羔数分别比CC基因型的多0.99(P〈0.01)和0.87只(P〈0.01)。济宁青山羊在P1、P5两个座位上都处于哈迪温伯格平衡状态(P1座位:χ^2=1.66,P=0.437;P5座位:χ^2=1.22,P=0.544)。
In present study, the luteinizing hormone beta-subunit (LHβ) gene was selected as a candidate gene for the high prolificacy in Jining Grey goats. According to the sequence of ovine LHβ gene, six pairs of primers were designed to detect single nucleotide polymorphisms in the 5' regulatory region, exon 1, exon 2 and exon 3 of ovine LHβ gene in high prolificacy breed (Jining Grey goat), medium prolificacy breed (Boer goat) and low prolificacy breeds (Liaoning Cashmere and Angora goats) by PCR-SSCP method. The results showed that the similarity in nucleotide sequence of goat and ovine LHβ gene was 98%, and only the products amplified by primers P1 and P5 displayed polymorphisms. For primer P1, three genotypes (AA, AB and BB) were detected in all four goat breeds. Sequencing revealed that two nucleotide substitutions (202C→A and 210C→T) at the 5'regulatory region of LHβgene in the BB genotype compared with the AA genotype. The differences of the litter size between different genotypes were nonsignificant in Jining Grey goats(P〉0.05). For primer P5, three genotypes (CC, CD and DD) were detected in Jining Grey goats, two genotypes (CC and CD) were detected in Liaoning Cashmere goats, and only one genotype CC was detected in Boer and Angora goats. Sequencing revealed one single nucleotide mutation (1124C→T) at exon 2 of LHβgene in the DD genotype compared with the CC genotype. Genotype frequency of CC, CD and DD was 0.38, 0.44 and 0.18 in Jining Grey goats, respectively. The Jining Grey does with genotype DD or CD had 0.99 (P〈0.01) or 0.87 (P〈 0.01) kids more than those with genotype CC. The fitness tests showed that the Jining Grey goat population was in Hardy-Weinberg equilibrium (P1 locus: Х^2 = 1.66, P= 0. 437 ; P5 locus: Х^2= 1.22, P=0. 544).