Research On Mevalonate Pathway Genes Of Chinese White Pine Beetle(Dendroctonus Armandi)

The Chinese white pine beetle(Dendroctonus armandi Tsai and Li) is arguably the most destructive forest insect in the Qinling Mountains, which attacks healthy Chinese white pine(Pinus armandi Franch) trees aged over 30 years, can kill large area living P. armandi and bring about huge economic losses. Biochemical studies have demonstrated that the mevalonate pathway is essential for de novo pheromone biosynthesis in several species of Scolytidae. The biosynthesisis regulated by juvenile hormone III, which is a sesquiterpenoid product of the mevalonate pathway. Therefore, study on expression of the related genes of mevalonate pathway amongst development stages, tissue distribution, fed on phloem of Pinus armandi and topically appliedjuvenile hormone(JH) III, it may contribute to understanding the relationship between D. armandi and its semiochemical production.Reasearch according to cloning, RACE and qRT-PCR technique to study quantitative expression of D. armandi DaHMG-R, DaHMG-S, DaG/FPPS and DaIDI genes at different development stages, tissues, JH III and feeding treatments. The main results are as follows: 1. We got four genes of mevalonate pathway, the full-length of the four genes were 3014 bp, 2018 bp, 1573 bp and 936 bp, repectively. By the comparison of amino acid sequence, we confirmed the four genes belong to HMG-CoA_reductase, cond_enzymes, Isoprenoid_ Biosyn_C1 and Nudix_Hydrolose families, respectively, and they were given the name of DaHMG-R、DaHMG-S、DaG/FPPS and DaIDI. 2. DaHMG-R, DaHMG-S, DaG/FPPS and DaIDI encoded 849, 457, 429 and 252 amino acids, respectively. Molecular weight and theoretical isoelectric point of DaHMG-R were 93.43 KDa and 7.52, respectively. Molecular weight and theoretical isoelectric point of DaHMG-S were 50.72 KDa and 6.97, respectively. Molecular weight and theoretical isoelectric point of DaIDI were 49.55 KDa and 8.32, respectively. Molecular weight and theoretical isoelectric point of DaG/FPPS were 28.99 KDa and 6.42, respectively. 3. The development stage of DaHMG-R, DaHMG-S, DaG/FPPS and DaIDI transcript levels was determined using qRT-PCR. Significant differences were found between different levels(p<0.05), and the stage of emerge adult had the highest expression of the four genes. Statistical significant differences were not found between sexes in teneral adults, but differences in transcript expression levels were found between sexes in emerge adults. 4. The tissue distribution of DaHMG-R, DaHMG-S, DaGPPS/FPPS and DaIDI mRNA in JH III- or acetone-treated(control) males and females was determined by qRT-PCR after a 20 h time period. Significant differences were found between different tissues(p<0.05), and the anterior midgut had the highest expression of the four genes. In acetone-treated males and females, statistical significant differences were found between tissue distribution, and expression of four genes was induced by JH III, especially in anterior midgut and hindgut. 5. DaHMG-R, DaHMG-S, DaGPPS/FPPS and DaIDI mRNA in time and dose of JH III-treated(control) males and females was determined by qRT-PCR. The mRNA levels increased while the dose and time was increased, when reached a max value, the mRNA levels began to decrease. The mRNA levels of males were higher than females at every time and dose points studied. 6. Analysed the mRNA levels of DaHMG-R, DaHMG-S, DaG/FPPS and DaIDI genes in feeding males and females. The mRNA levels increased while the time increased, when reached a max value, the mRNA levels began to decrease. Statistical significant differences were found in feeding males and females(p<0.05), and the mRNA levels of males were higher than females at every time points studied.