关键词: Y染色体, 绵羊, 单核苷酸多态性

Abstract: 【Objective】The male specific region of the mammalian Y chromosome (MSY)does not recombine with X chromosome during meiosis process, which is an important genetic resource for analyzing paternal genetic diversity due to its strict father to son inherited character. In addition, most of genes on MSY exclusively or predominantly express in the testis, indicating they may play essential roles in spermatogenesis and male reproduction. Since it is extremely difficult to sequence of entire Y chromosome, many species have very few Y chromosome sequences. Therefore, the current study was conducted to select ovine Y chromosome specific primers and Y-SNP based on the previous primer information used in bovidae species, and to compare the Y-fragment sequences similarity among sheep, bovine, goat, yak and bharal. Meanwhile, the Y-SNP was associated with sheep scrotal circumference to supply scientific basis for constructing ovine Y-haplotypes, identifying molecular markers for embryo sex and male reproductive traits in the future.【Method】Based on the investigation of available references about bovidae Y chromosome, 29 pairs of Y-primers reported in cattle, yak, goat were selected to amplify rams DNA using ewes DNA and ddH₂O as negative controls. Subsequently, the Y-SNPs within ovine Y-specific fragment identified of different sheep breeds were investigated by DNA sequencing of DNA pooling and PCR-RFLP methods, including Suffolk sheep (n=146),White Suffolk sheep (n=91), East Friesian sheep (n=6), Texel sheep (n=72), South African Mutton Merino (n=17), Dorper sheep (n=32), Hu sheep (n=55), Tibetan sheep (n=34), Tan sheep (n=43), and bharal (n=14). Chromas and DNASTAR were used to analyze the results of DNA-pool sequencing, and DNAman was used for homology analysis of yak, goat, cattle and bharal. Meanwhile, the correlation analysis between the SRY11 gene fragment polymorphisms and the testis size was performed by SPSS 19.0.【Result】The results showed that 6 out of 29 pairs of primers analyzed were ovine Y-specific, which could amplify ZFY3, SRY6, USP9Y, UTY, SRY11 and ZFY6 fragments, respectively. However, 17 pairs of primers failed to show amplification bands, and 6 pairs of primers showed amplified bands in the DNA of the ewes. The similarity of them among sheep, bharal, cattle, goat and yak ranged from 81.51% to 98.84%. In addition, a Y-SNP (G＞A) with in SRY11 fragment was first identified in the Suffolk and white Suffolk sheep. According to RCR-RFLP analysis, two genotypes (AA, GG) were detected in the Suffolk sheep and white Suffolk sheep, while only the GG genotype was found in the other seven sheep breeds. The genotypic frequencies of the GG and AA were 0.747 and 0.253 in White Suffolk sheep, respectively, while they were 0.986 and 0.014 in Suffolk sheep, indicating the dominant genotype was GG genotype in White Suffolk sheep and Suffolk sheep. Association analysis suggested that the testis size of the GG genotype was significantly higher than those of the AA genotype in the white Suffolk sheep population (P=0.029).【Conclusion】In this study, six pairs of ovine Y-specific primers were identified, and the Y-linked fragment identified in ovine showed a high similarity with cattle, goats and yaks, indicating certain conservation in the evolutionary process. In addition, a Y-SNP was found to be specific in White Suffolk sheep and Suffolk sheep, which was closely associated with the testis size of white Suffolk sheep.

Key words: Y chromosome, sheep, single nucleotide polymorphism (SNP)