一个新的家蚕kunitz类型CI基因的克隆和序列分析

Molecular Cloning and Sequence Analysis of a Novel Kunitz-type Chymotrypsin Inhibitor Gene of Bombyx mori

家蚕胰凝乳蛋白酶抑制剂(chymotrypsin inhibitor,CI)的多个编码基因位于多个染色体位点上,是家蚕CI具有丰富多态性的原因之一。根据Gene Bank中登录的家蚕CI基因Sci-sb的mRNA,设计引物克隆了一个家蚕kunitz类型的CI基因,命名为CI-fb。CI-fb基因有3个外显子,开放阅读框长度为255 bp,编码85个氨基酸残基,具有一个典型的kunitz结构域。CI-fb基因与其他已知的kunitz类型的CI基因相比,在基因序列和结构、蛋白序列和结构上高度相似,但在已知的CI基因家族中序列最短。RT-PCR分析表明,CI-fb基因在家蚕的脂肪体、精巢、卵巢、马氏管、气管和中肠6个组织中表达,而在丝腺、血细胞和体壁中没有表达,是一个组织特异性表达的基因。研究结果为进一步研究家蚕CI的多态性和功能提供了新的线索。

Chymotrypsin inhibitor （CI） encoding genes lie in multiple sites of chromosomes, which results in rich polymorphism of chymotrypsin inhibitor in Bombyx mori. A novel kunitz-type CI gene of silkworm, CI-fb, was cloned with the specific primers designed according to the mRNA of Sci-sb recorded in NCBI. The CI-fb contains three exons and a complete ORF of 255 bp coding a putative precursor protein of 85 amino acid residues with typical kunitz domain. The CI-fb is similar to the other known kunitz-type CI encoding genes in gene sequence and structure, protein sequence and structure, but its genome sequence is the shortest. By RT- PCR analysis, CI-fb was detected in six tissues including fat body, ovary, testis, Malpighian tubule, trachea and midgut, but not in silk grand, blood cell and skin, suggesting CI-fb is a tissue-specific expression gene. This result gives a new clue for studying the polymorphism and function of CIs of Bombyx mori.