Chongqing Key Laboratory of Entomology and Pest Control Engineering, College of Plant Protection, Southwest University, Chongqing 400716, China.
J Insect Sci. 2010;10:112. doi: 10.1673/031.010.11201.
In this study, the cDNA of Tetranychus cinnabarinus (Boisduval) (Acarina: Tetranychidae) HSP90 (designated TcHSP90) was cloned using a combination of the homology cloning and rapid amplification of cDNA ends (RACE) approaches. The full-length cDNA of TcHSP90 is 2595 bp, including a 5′-untranslated region (UTR) of 177 bp, 3′-UTR of 249 bp, and an open reading frame (ORF) of 2169 bp. The ORF encodes a polypeptide of 722 amino acids with a predicted molecular weight of 83.45 kDa and a theoretical isoelectric point of 4.81. There is an mRNA polyadenylation signal of ATTAAA at the positions 2558-2564. In addition, the expression pattern of TcHSP90 mRNA relative to that of beta-actin gene in the three stains of T. cinnabarinus (AbR, abamectin-resistant strain; HR, heat-resistant strain; SS, the susceptible strain) were examined by using fluorescent real time quantitative PCR after the impact of abamectin, high and low temperature, respectively. The results showed that under the normal condition, the mRNA level of TcHSP90 was 1.64 and 1.29-fold higher in the AbR and HR than in SS, respectively. After 8 h treatment with abamectin, the TcHSP90 mRNA levels of SS, AbR, and HR were 1.25, 1.87, and 2.05-fold higher than those of their untreated controls, respectively. The TcHSP90 mRNA levels of SS, AbR, and HR were also significantly increased after being induced at 40 degrees C for 1 h, and they were 3.76, 3.42, and 3.79-fold higher than those of their untreated controls, respectively. The mRNA level of TcHSP90 was also significantly increased after being induced at 4 degrees C for 1 h. These results suggest that TcHSP90 might be involved in the abamectin and extreme temperature resistance or tolerance.
本研究采用同源克隆和快速扩增 cDNA 末端（RACE）技术克隆了朱砂叶螨（Tetranychus cinnabarinus）（蜱螨目：叶螨科）HSP90（命名为 TcHSP90）的 cDNA。TcHSP90 的全长 cDNA 为 2595 bp，包括 177 bp 的 5′非翻译区（UTR）、249 bp 的 3′UTR 和 2169 bp 的开放阅读框（ORF）。ORF 编码一个 722 个氨基酸的多肽，预测分子量为 83.45 kDa，理论等电点为 4.81。在 2558-2564 位有一个 mRNA 多聚腺苷酸化信号 ATTAAA。此外，在用阿维菌素、高低温分别处理后，通过荧光实时定量 PCR 检测了 T. cinnabarinus 三个品系（AbR，抗阿维菌素品系；HR，耐高温品系；SS，敏感品系）中 TcHSP90 mRNA 相对于β-肌动蛋白基因的表达模式。结果表明，在正常条件下，AbR 和 HR 中的 TcHSP90 mRNA 水平分别比 SS 高 1.64 倍和 1.29 倍。SS、AbR 和 HR 在用阿维菌素处理 8 h 后，其 TcHSP90 mRNA 水平分别比未处理对照组高 1.25、1.87 和 2.05 倍。SS、AbR 和 HR 在 40℃诱导 1 h 后，TcHSP90 mRNA 水平也显著升高，分别比未处理对照组高 3.76、3.42 和 3.79 倍。在 4℃诱导 1 h 后，TcHSP90 mRNA 水平也显著升高。这些结果表明，TcHSP90 可能参与了阿维菌素和极端温度的抗性或耐受性。
