Ahlawat S, Chopra M, Jaiswal L, Sharma R, Arora R, Brahma B, Lal S V, De S
National Bureau of Animal Genetic Resources, Karnal, 132001, India.
National Dairy Research Institute, Karnal, 132001, India.
Mol Cell Probes. 2016 Apr;30(2):66-73. doi: 10.1016/j.mcp.2016.03.001. Epub 2016 Mar 3.
Disrupted meiotic cDNA1 (DMC1) recombinase plays a pivotal role in homology search and strand exchange reactions during meiotic homologous recombination. In the present study, full length coding sequence of DMC1 gene was sequence characterized for the first time from four ruminant species (cattle, buffalo, sheep and goat) and phylogenetic relationship of ruminant DMC1 with other eukaryotes was analyzed. DMC1 gene encodes a putative protein of 340 amino acids in cattle, sheep and buffalo and 341 amino acids in goat. A high degree of evolutionary conservation at both nucleotide and amino acid level was observed for the four ruminant orthologs. In cattle and sheep, novel alternatively spliced mRNAs with skipping of exons 7 and 8 (Transcript variant 1, TV1) were isolated in addition to the full length (FL) transcript. Novel transcript variants with partial skipping of exon 7 and complete skipping of exon 8 (Transcript variant 2, TV2) were found in sheep and goat. The presence of these variants was validated by amplifying cDNA isolated from testis tissue of ruminants using two oligonucleotides flanking the deleted region. To accurately estimate their relative proportions, real-time PCR was performed using primers specific for each variant. Expression level of DMC1-FL was significantly higher than that of TV1 in cattle and TV2 in goat (P < 0.05). Relative ratio for expression of DMC1-FL: TV1: TV2 in sheep was 6.78: 1.43: 1. In-silico analysis revealed presence of splice variants of DMC1 gene across other mammalian species underpinning the role of alternative splicing in functional innovation.
减数分裂破坏cDNA1（DMC1）重组酶在减数分裂同源重组过程中的同源性搜索和链交换反应中起关键作用。在本研究中，首次对四种反刍动物（牛、水牛、绵羊和山羊）的DMC1基因全长编码序列进行了序列特征分析，并分析了反刍动物DMC1与其他真核生物的系统发育关系。牛、绵羊和水牛的DMC1基因编码一个推定的340个氨基酸的蛋白质，山羊的DMC1基因编码一个推定的341个氨基酸的蛋白质。观察到这四个反刍动物直系同源物在核苷酸和氨基酸水平上都具有高度的进化保守性。在牛和绵羊中，除了全长（FL）转录本外，还分离出了外显子7和8跳跃的新型可变剪接mRNA（转录本变体1，TV1）。在绵羊和山羊中发现了外显子7部分跳跃和外显子8完全跳跃的新型转录本变体（转录本变体2，TV2）。使用位于缺失区域两侧的两种寡核苷酸扩增从反刍动物睾丸组织分离的cDNA，验证了这些变体的存在。为了准确估计它们的相对比例，使用针对每个变体的特异性引物进行了实时PCR。在牛中，DMC1-FL的表达水平显著高于TV1，在山羊中显著高于TV2（P < 0.05）。绵羊中DMC1-FL: TV1: TV2的相对表达比例为6.78: 1.43: 1。电子分析揭示了DMC1基因的剪接变体在其他哺乳动物物种中的存在，这支持了可变剪接在功能创新中的作用。
