绵羊胚胎性别鉴定试剂盒的研究

Establishment of PCR Sex Identification Kit of Sheep Embryos

根据绵羊Y-染色体的特异序列和常染色序列分别设计了确定公羊Y-染色体特异序列的3对特异性引物和内标基因的4对特异性引物。单重PCR扩增绵羊基因组DNA,筛选出了3对Y-染色体特异引物和3对羊DNA特异内标引物。将不同的绵羊Y-染色体特异引物与内标引物组合,利用多重PCR扩增绵羊基因组DNA,筛选出了1个可用于羊早期胚胎性别鉴定的PCR引物组合:A0/C1。按照最优PCR扩增DNA条件配制了绵羊PCR性别鉴定试剂盒并成功应用于绵羊血液、已知性别的绵羊成纤维细胞和胚胎,表明本研究建立的体系完全可用于绵羊早期的胚胎性别鉴定。

3 pairs of sheep male-specific PCR primers were designed based on the sequences of sheep Y-specific repeat sequence, and 4 pairs of sheep-specific PCR primers were designed as internal control primers, After PCR amplification of sheep genomic DNA with different primers, 3 pairs of sheep male-specific primers and 3 pairs of sheep-specific primers could work very well. A0/C1 combination of sheep male- speeifie PCR primers and internal eontrol primers were obtained for sexing sheep pre-implantation embryo after multiplex PCR amplifications of sheep genomie DNA. On basis of the optimal DNA PCR conditions for SRY primers, the PCR sex determination kit was developed and tested sheep genomie DNA of blood, fibroblasts and embryos. Therefore, the system constructed in this experiment could be used completely to determine the sex of sheep early embryos.