RNAi技术介导舞毒蛾热激蛋白Hsp40基因功能分析

Functional Analysis of Hsp40 Gene in Lymantria dispar Mediated RNAi Technology

[目的]测定Hsp40基因沉默对舞毒蛾生长发育及Hsp40基因表达量的影响。[方法]体外合成双链RNA(dsRNA),并将dsRNA通过微注射入舞毒蛾3龄幼虫体内,测定Hsp40基因沉默对舞毒蛾生长发育及Hsp40基因表达量的影响。[结果]分别注射ddH2O、dsRNAGFP和dsRNAHsp40后8 d,dsRNAGFP处理组舞毒蛾幼虫相对取食量显著高于ddH2O和dsRNAHsp40处理组(P<0.05);但3种处理对舞毒蛾幼虫的相对生长率、食物利用率、食物转化率、近似消化率方面均无显著性差异。注射后4 d,ddH2O处理组的舞毒蛾幼虫体重累计增长率最大,其次是dsRNAHsp40处理组,dsRNAGFP处理组的体重累计增长率最小,这与4 d的幼虫鲜重一致;其余时间点,dsRNAHsp40处理组的体重累计增长率均大于ddH2O和dsRNAGFP处理组。将1μl(1μg/μl)的dsRNA注射入舞毒蛾幼虫体内,6~48h Hsp40基因表达量显著下降,96 h基因表达量上调。[结论]该研究为进一步利用沉默舞毒蛾Hsp40基因在害虫防治中的应用提供理论依据。

[ Objective ] The aim was to measure the effects of Hsp40 gent silencing on growth and Hsp40 gene expression of L. d/spar larvae. [Method] The 1 txg/l^l of double-stranded RNA （dsRNA） in vitro synthesized,was microinjected into 3rd instar Lymararia dispar larvae. The effects of Hsp40 gene silencing on growth and Hsp40 gene expression of L. d/spar larvae were measured. [ Result] The results showed the relative consumption rate （RCR） of L. dispar larvae microinjected by dsRNAGFP was higher than those microinjected by ddH20 and dsRNAHslMO at 8 d time point. However, relative growth ratio （RGR）, efficiency of conversion of ingested food （ECI）, approximate digestibility （AD）, efficien- cy of conversion of digested food （ECD） of L. d/spar larvae among three treatments were no significant differences. After 4 d of microinjection, the weight cumulative growth rates of L. dispar larvae were decreasing order of ddH2O, dsRNAHsp40 and dsRNAGFP, which were consistent with the larvae weight. At the rest of time points, weight cumulative growth rate of dsRNAHsp40 treatment group were greater than other treat- ment groups. Mter microinjection 1 μg/μl of dsRNA into the L d/spar larvae ranged from 6 h to 48 h, Hsp40 gene expressions were significantly decreased ,while those increased at 96 h. [ Conclusion] These results provided a theoretical basis for further silencing Hsp40 gene of L d/spar into pest control.