苜蓿实夜蛾几丁质脱乙酰基酶基因cDNA序列的克隆与序列分析

摘要

Chitin deacetylase (CDA) plays an important role in insect chitin metabolism. Total RNA was isolated from the fifth instar larvae of the marbled clover, Heliothis viriplaca Hüfnagel. A H. viriplaca CDA cDNA sequence was cloned by RT-PCR and rapid amplification of cDNA ends (RACE). The CDA cDNA was 1 984 bp in length, which contained an open reading frame of 1 626 bp and encoded a polypeptide of 541 aa residues with a predicted molecular weight of about 61.86 ku. The deduced amino acid sequence contained a polysaceharide deacetylase-like catalytic domain, a chitin-binding peritrophin-A domain (ChBD) and a low-density lipoprotein receptor class A domain (LDLa). The H. viriplaca CDA cDNA and the deduced amino acid sequence shared different levels of identity with those from other insects. The amino acid sequence identity between H. viriplaca CDA and H. armigera CDA (GQ411189) was 98％, but only 36％-38％ between H. viriplaca CDA and Trichoplusia ni CDA (AY966402), Mamestra configurata CDA (U660852) and H. armigera CDA (GQ411190GQ411191EF600051), respectively. This H. viriplaca CDA belonged to one of the two types of CDA. The cDNA sequence has been deposited in GenBank (GU188855).%几丁质脱乙酰基酶(CDA)在昆虫几丁质代谢中具有重要作用.本研究以苜蓿实夜蛾5龄幼虫虫体为材料提取总RNA,利用RT-PCR和RACE技术,扩增得到该虫CDA基因的cDNA序列.该序列含有1 984个碱基,包括1个1 626个碱基的开放阅读框,编码一个含541个氨基酸的蛋白,分子量约为61.86 ku.克隆基因推导的氨基酸序列具有几丁质脱乙酰基酶典型的3个功能区,分别是几丁质结合区(47～103),催化区(199～355),低密度脂蛋白受体区(124～158).序列比对表明,克隆的苜蓿实夜蛾CDA与其他昆虫的CDA在核苷酸和氨基酸水平上的一致性存在较大差异.苜蓿实夜蛾CDA的氨基酸序列与棉铃虫CDA(GQ411189)一致性达到98%,而与另外3种昆虫粉纹夜蛾CDA(AY966402)、蓓带夜蛾CDA(EU660852)和棉铃虫CDA(GQ411190GQ411191EF600051)氨基酸序列间的一致性只有36%～38%,属于两类CDA类群中的一类.基因的cDNA序列已经登录GenBank并获得登录号GU1188855.