Zhang Ranran, Gao Guanqun, Chen Hui
College of Forestry, Northwest A&F University, Yangling, Shaanxi 712100, China.
Sci Rep. 2016 Mar 16;6:23136. doi: 10.1038/srep23136.
In this study, a polymerase chain reaction (PCR) based on homology genes of Orco was utilized to identify DarmOrco, which is essential for olfaction in D. armandi. The results showed that DarmOrco shares significant sequence homology with Orco proteins had known in other insects. Quantitative real-time PCR (qRT-PCR) analysis suggested that DarmOrco was abundantly expressed in adult D. armandi; by contrast, DarmOrco showed trace amounts of expression level in other stages. Of different tissues, DarmOrco expression level was the highest in the antennae. In order to understand the functional significance of Orco, we injected siRNA of DarmOrco into the conjunctivum between the second and third abdominal segments, and evaluated its expression after siRNA injected for 24 h, 48 h and 72 h. The results of qRT-PCR demonstrated that the reduction of mRNA expression level was significant (80%) in DarmOrco siRNA-treated D. armandi than in water-injected and non-injected controls. The electroantennogram responses of females and males to 11 major volatiles of its host, were also reduced (3068% for females; 16~70% for males) in siRNA-treated D. armandi compared with the controls. These results suggest that DarmOrco is crucial in mediating odorant perception.
在本研究中,利用基于气味受体共同蛋白(Orco)同源基因的聚合酶链式反应(PCR)来鉴定华山松大小蠹(Dendroctonus armandi)中的DarmOrco,它对华山松大小蠹的嗅觉至关重要。结果表明,DarmOrco与其他昆虫中已知的Orco蛋白具有显著的序列同源性。实时定量PCR(qRT-PCR)分析表明,DarmOrco在华山松大小蠹成虫中大量表达;相比之下,DarmOrco在其他发育阶段的表达量极低。在不同组织中,DarmOrco在触角中的表达水平最高。为了了解Orco的功能意义,我们将DarmOrco的siRNA注射到第二和第三腹节之间的结膜中,并在注射siRNA 24小时、48小时和72小时后评估其表达情况。qRT-PCR结果表明,与注射水和未注射的对照相比,经DarmOrco siRNA处理的华山松大小蠹中mRNA表达水平显著降低(约80%)。与对照相比,经siRNA处理的华山松大小蠹的雌虫和雄虫对其寄主的11种主要挥发物的触角电位反应也降低了(雌虫为30%68%;雄虫为16%70%)。这些结果表明,DarmOrco在介导气味感知中起关键作用。