小菜蛾GluCl受体α亚基cDNA基因克隆和序列分析

Cloning and sequence analysis of cDNA of GluCl receptor alpha gene from the diamondback moth,Plutella xylostella

利用RT-PCR和RACE技术对小菜蛾[Plutella xylostella(L.)]4龄幼虫谷氨酸门控的氯离子通道(GluCl)受体cDNA全长进行了克隆和序列分析。通过设计简并性上、下游引物扩增出小菜蛾GluCl受体α亚基基因192bp的cDNA片段,根据得到的片段序列设计3′和5′特异性引物采用RACE技术进行3′和5′末端的克隆,获得了小菜蛾GluCl受体的cDNA的完整序列,GenBank登录号为GQ221939。该基因全长2144bp,其开放阅读框(ORF)1344bp,编码447个氨基酸。相似性分析表明:它与果蝇和赤拟谷盗的相似性均为73%,与埃及伊蚊的相似性为75%,与东亚飞蝗的相似性为79%;氨基酸分析后显示它具有GluClα亚基的典型特征,表明克隆的cDNA序列是小菜蛾GluClα亚基基因的序列。

One full-length eDNA sequence of the GluCl receptor alpha gene of the fourth instars of the diamondback moth, Plutella xylostella （L.） was amplified, cloned, sequenced and analyzed by using RT-PCR and RACE method. One fragment of 192 bp cDNA of the GluCl receptor alpha gene was obtained by using degenerate forward and reverse primers. Then the 5′-and 3′-end specific primers were designed according to the fragment sequence for the 5＇-and 3＇-RACE. A 2 144 bp full-length cDNA of GluC1 receptor was obtained. This gene was deposited in GenBank （accession no. GO221939）. It has an open reading frame of 1 344 nucleotides, encoding 447 amino acids residues. Similarity analysis showed that the cloned cDNA sequence had 73% identity with the published sequences of Drosophila rnelanogaster and Tribolium castaneurn, 75% and 79% identities with those of Aedes aegypti and Locusta migratoria. Typical features of GluC1 subunits as in other members of the Cys-loop lig- and-gated ion channel superfamily were found through protein analysis. The results indicated that the cDNA sequence was GluClα receptor gene.