【摘要】 【目的】本研究旨在获得家蚕Bombyx mori Bcl-2家族同源基因,并分析其在不同组织和发育阶段的时空表达模式及功能。【方法】用c DNA末端快速扩增方法(rapid amplification of c DNA ends,RACE)克隆家蚕Bcl-2家族基因Bm Buffy,利用SSR分子标记连锁分析确定其染色体定位。同时,用RT-PCR和q PCR技术分析该基因在家蚕幼虫及变态期间不同组织中的表达。【结果】克隆获得全长家蚕Bcl-2家族同源基因Bm Buffy,证明该基因位于第4号染色体上,开放阅读框长879 bp,编码292 aa,预测其分子量大小为32.4 k Da,等电点为9.94,且第130-231位氨基酸之间存在1个Bcl-2_like Superfamily结构域。系统进化发育树表明,其与黑腹果蝇Drosophila melanogaster的Dm Buffy关系最近,氨基酸序列一致性为27%。Bm Buffy在幼虫组织中的表达结果显示,其在马氏管中表达量最高,而且在不同组织变态期的关键时间点均有明显变化。【结论】家蚕Bm Buffy具有Bcl-2家族典型结构域,Bm Buffy定位于第4号染色体上,Bm Buffy在家蚕变态期的组织生理变化中起到一定作用。本文为进一步研究家蚕Bcl-2家族基因的功能奠定基础。更多还原

【Abstract】 【Aim】This study aims to clone a Bcl-2 family member gene from the silkworm,Bombyx mori,and explore its expression profiles and roles in different larval tissues and developmental stages of the moth. 【Methods】Bm Buffy was cloned using rapid amplification of c DNA ends( RACE) method.RT-PCR and q PCR were used to detect the gene expression profiles. The chromosome localization of this gene was conducted using SSR markers. 【Results】The Bcl-2 family member gene cloned in B. mori was named Bm Buffy,which is located on chromosome No. 4,coding a protein of 292 aa with an estimated molecular weight of 57. 8 k Da and p I of 9. 94. Its encoded protein also possesses a Bcl-2 like superfamily domain between 130- 231 residues. Phylogenetic analysis revealed that Bm Buffy is homologous to Dm Buffy of Drosophila melanogaster with 27% amino acid sequence identity. The expression profiles in different larval tissues showed that Bm Buffy had the highest expression level in Malpighian tubules. There was significant variation in the transcription level of Bm Buffy in different developmental stages including metamorphosis. 【Conclusion 】 Bm Buffy has the typical Bcl-2 family domain,Bm Buffy is located on chromosone No. 4,and Bm Buffy might play a role in the alteration of tissue physiology during metamorphosis.This study lays a foundation for further study on the function of Bcl-2 family genes in B. mori.更多还原