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为明确vasa基因在中华卵索线虫Ovomermis sinensis生殖系发育以及原始生殖细胞增殖过程中的作用,采用同源克隆与cDNA末端快速扩增技术(RACE),从中华卵索线虫中克隆得到vasa基因全长cDNA序列(3053bp),编码813个氨基酸,具有DEAD-box家族蛋白全部9个保守基序。经同源比对与系统进化树分析,确定其属于DEAD-box家族的VASA亚家族成员。利用反转录PCR(reverse transcription PCR,RT-PCR)以及实时荧光定量PCR(real time PCR)分析,vasamRNA在中华卵索线虫性腺中专一性表达,其它部位如头、尾、体壁均未检测到vasa表达信号,且在生殖腺发育初期表达量最高。此外,vasa基因转录本在中华卵索线虫胚胎发育期卵中表达量较高,证实其为母源性基因,调控中华卵索线虫胚胎发育的过程。
In order to clarify the role of vasa gene in the germination of Ovomermis sinensis and the proliferation of primordial germ cells, the homologous cloning and rapid amplification of cDNA ends (RACE) were used to clone the vasa gene from Ovomermis sinensis The long cDNA sequence (3053bp) encoded 813 amino acids with all nine conserved motifs of the DEAD-box family of proteins. After homologous comparison and phylogenetic tree analysis, the members of VASA subfamily belonging to DEAD-box family were identified. Using reverse transcription PCR (RT-PCR) and real-time PCR analysis, vasamRNA was exclusively expressed in the gonads of C. sinensis and all other sites, such as head, tail and body wall Vasa expression signal was detected, and the highest expression in the early stage of gonadal development. In addition, the vasa gene transcripts were highly expressed in the eggs of C. sinensis during embryogenesis, confirming that it is a maternal gene that regulates the embryonic development of C. sinensis.