舞毒蛾DnaJ1基因克隆分析及对甲萘威胁迫响应

Dna J蛋白是Dna K/Hsp70的辅助分子伴侣,通过调节Hsp70的ATPase活性来影响蛋白复合体的合成与组装。为明确舞毒蛾Lymantria dispar的LdDnaJ1基因特性及对杀虫剂甲萘威的胁迫响应,通过克隆LdDnaJ1全长基因并运用实时荧光定量RT-PCR技术测定了甲萘威对其LdDnaJ1基因表达量的影响。结果表明,舞毒蛾Ld Dna J1全长基因开放阅读框为1 062 bp,编码353个氨基酸,分子质量为39.91 kD,理论等电点为5.65;舞毒蛾Dna J与柑橘凤蝶Papilio xuthus Dna J亲缘关系较近。甲萘威对舞毒蛾2龄幼虫24 h和48 h的致死中浓度LC50分别为74.04 mg/L和31.48mg/L。低剂量(LC_5、LC_(10)和LC_(30))甲萘威胁迫下,舞毒蛾2龄幼虫Ld Dna J1基因表达量均下调,LC_(30)甲萘威处理后72 h时LdDnaJ1基因表达量最低,仅为对照的15.70%。表明甲萘威可抑制舞毒蛾LdDnaJ1基因的表达,且呈现明显的时间和剂量效应。

Cloning analysis of LdDnaJ1 in gypsy moth Lymantria dispar and its response to carbaryl stress

DnaJ protein is a molecular chaperone of DnaK/Hsp70 mediating protein complex synthesis and assembly by regulating the ATPase activity of Hsp70 protein. To investigate the characteristics of LdDnaJ1 in gypsy moth Lymantria dispar (Linnaeus) and its response to carbaryl stress, the full-length cDNA of LdDnaJ1 was cloned and the effects of carbaryl on LdDnaJ1 gene expression were measured by using real-time RT-PCR technology. The results showed the open reading frame of LdDnaJ1 was 1062 bp, encoding 353 amino acid residues with a molecular mass of 39.91 kD and theoretical pI of 5.65. Phylogenetic analysis of DnaJ proteins showed that LdDnaJ1 clustered into a group with Papilio xuthus. The 24-h and 48-h LC₅₀ of carbaryl for 2nd instar L. dispar larvae was 74.04 mg/L and 31.48 mg/L, respectively. Under the low doses (LC₅, LC₁₀ and LC₃₀) of carbaryl stress, the expression of LdDnaJ1 in 2nd instar L. dispar larvae was obviously downregulated during a 72 h-period. The expression of LdDnaJ1 was the lowest at the time point of 72 h with only 15.7% of that in the control under LC₃₀ dose of carbaryl stress. These results indicated that the expression of LdDnaJ1 was inhibited by carbaryl and responded to carbaryl in a concentration- and time-dependent way.