云南切梢小蠹热激蛋白40基因的克隆与序列分析

目的]克隆云南切梢小蠹(Tomicus yunnanensis)热激蛋白40基因,并进行序列分析,得到该基因的特征。方法]采用RACE克隆技术,获得云南切梢小蠹热激蛋白40基因,并对该基因进行分析。结果]试验获得的基因序列长为1 205 bp,开放阅读框1173 bp,编码氨基酸序列390个,编码蛋白质的预测分子量为43.56 ku,等电点为7.35。该基因蛋白序列具有1个Amidation位点,2个天冬酰胺N末端糖基化位点,2个CAMP和CGMP依赖性蛋白激酶磷酸化位点,3个酪蛋白激酶Ⅱ磷酸化位点,7个N末端十四烷基化位点,4个蛋白激酶C磷酸化位点,1个RGD细胞附着序列,1个DNAJ-1结构域等热激蛋白40所具有的典型特征。结论]通过同源性比对分析发现,云南切梢小蠹热激蛋白40基因与家蚕(Bombyx mori)、赤拟谷盗(Tribolium castaneum)、美洲斑潜蝇(Liriomyza sativae)和东亚飞蝗(Locusta migratoria)的热激蛋白40基因在氨基酸水平上一致性为20%~30%,它们的相似性均不高。虽然不同昆虫Hsp40在进化过程中具有高度的保守性,编码序列变异比较低,但是结构域差异与生物体适应外界环境有一定的关系。

Molecular Cloning and Sequence Analysis of the Heat Shock Protein 40 Gene in Tomicus yunnanensis

Objective] To clone the gene of heat shock protein 40 in Tomicus yunnanensis and get the gene characteristics by sequence analysis.Method] The gene of heat shock protein 40 in T.yunnanensis was obtained and analyzed by using rapid amplification of cDNA ends (RACE) method.Result]This gene was 1 205 bp in length with opening reading frame (ORF) of 1 173 bp.The deduced amino acid sequence of T.yunnanensis Hsp40 revealed a protein of 390 amino acids.Its predicted molecular weight and isoelectric point were 43.56 ku and 7.35.It has several conservation amino acid sites including one Amidation site,two N-glycosylation sites,two CAMP and CGMP dependent protein kinase phosphorylation sites,three Casein kinase Ⅱ phosphorylation sites,seven N-myristoylation sites,four Protein kinase C phosphorylation sites,one RGD Cell attachment sequence,and one DNAJ-1 Cell attachment sequence.Conclusion] It shared 20％-30％ amino acid identity with the Hsp40 of Bombyx mori,Tribolium castaneum,Liriomyza sativae and Locusta migratoria.Their similarity is not high.Although the Hsp40 of different insects is highly conserved in the evolution process,the variation of the coding sequence is relatively low,but there has a certain relationship between the structural domain differences and the organism that adapting to the external environment.