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根据斑马鱼GDF9、BMP15基因的保守序列设计引物,采用RT-PCR方法扩增获得日本白鲫GDF9、BMP15基因片段。日本白鲫GDF9基因片段长778 bp,编码145个氨基酸;BMP15基因片段长811 bp,编码270个氨基酸。氨基酸序列分析表明日本白鲫GDF9与斑马鱼的同源性最高,为78%,与其他物种的同源性在58%～78%之间;日本白鲫BMP15与斑马鱼同源性最高,为80%,与其他物种的同源性在30%～80%之间。系统进化树分析显示,鱼类的GDF9基因独立聚成一支,其中日本白鲫的GDF9基因与斑马鱼的GDF9基因聚成一支;鱼类的BMP15基因独立聚成一支,其中日本白鲫的BMP15基因与斑马鱼的BMP15基因聚成一支。半定量PCR结果显示,BMP15 mRNA在脾脏、肝脏、鳃、脑、心脏、肌肉、肾脏、卵巢中均有表达,其中卵巢中表达量最高;GDF9 mRNA在肝脏、脑、心脏、肌肉、卵巢中均有表达,而在脾脏、鳃、肾脏中表达量极低或不表达,在卵巢中表达量最高,为进一步研究日本白鲫GDF9与BMP15基因的结构与功能奠定了基础。
Primers were designed according to the conserved sequences of GDF9 and BMP15 in zebrafish, and GDF9 and BMP15 gene fragments were amplified by RT-PCR. The length of GDF9 gene of Japanese white carp was 778 bp, encoding 145 amino acids. The length of BMP15 gene was 811 bp, encoding 270 amino acids. Amino acid sequence analysis showed that Japanese white crucian carp GDF9 had the highest homology with zebrafish (78%) and its homology with other species was 58% -78%. Japanese white crucian carp BMP15 had the highest homology with zebrafish 80%, with other species of homology between 30% ~ 80%. Phylogenetic tree analysis showed that the GDF9 gene of fish was independently aggregated, in which the GDF9 gene of Japanese white carp was clustered with the GDF9 gene of zebrafish; the BMP15 gene of fish was independently aggregated into one, and the BMP15 gene of Japanese white carp BMP15 gene and zebrafish together. Semi-quantitative PCR results showed that BMP15 mRNA was expressed in spleen, liver, gill, brain, heart, muscle, kidney and ovary, with the highest expression in ovary; GDF9 mRNA in liver, brain, heart, muscle and ovary The expression levels of GDF9 and BMP15 in Japanese white carp (Carassius auratus gibelio) are very low or not expressed in the spleen, gill and kidney, and they are the highest in the ovary.