Key Laboratory of Entomology and Pest Control Engineering, College of Plant Protection, Southwest University, Chongqing 400716, PR China.
J Insect Sci. 2010;10:208. doi: 10.1673/031.010.20801.
Reference genes are used as internal controls in gene expression studies, but their expression levels vary according to tissue types and experimental treatments. Quantitative real-time PCR (qPCR) is the most sensitive technique for transcript quantification provided that gene transcription patterns are normalized to an evaluated reference gene. In this study, the suitability of eight commonly used genes (β-actin, 5.8SrRNA, α-TUB, GAPDH, RPL13a, RPS18, TBP, SDHA) were cloned and investigated to find the most stable candidates for normalizing real-time PCR data generated from the four different strains (abamectin-resistant, fenpropathrin-resistant, omethoate-resistant, and susceptible strains) and different developmental stages (eggs, protonymphs, nymphs, and adults) of carmine spider mite, Tetranychus cinnabarinus (Boisduval) (Acarina: Tetranychidae). The stability of gene expression was assessed using two different analysis programs, geNorm and NormFinder. Using these analyses, RPS18 and 5.8SrRNA had the most stable expression regardless of the four different strains, whereas RPS18 and α-TUB were expressed most stably in different developmental stages.
内参基因被用作基因表达研究的内部对照，但它们的表达水平会根据组织类型和实验处理而变化。定量实时 PCR（qPCR）是一种最敏感的转录本定量技术，前提是将基因转录模式归一化为经过评估的内参基因。在这项研究中，克隆并研究了 8 个常用基因（β-肌动蛋白、5.8S rRNA、α-TUB、GAPDH、RPL13a、RPS18、TBP、SDHA），以找到最适合归一化胭脂红蜘蛛，Tetranychus cinnabarinus（Boisduval）（Acarina：Tetranychidae）四个不同品系（阿维菌素抗性、甲氰菊酯抗性、氧乐果抗性和敏感品系）和不同发育阶段（卵、前若虫、若虫和成虫）实时 PCR 数据的最稳定候选基因。使用两种不同的分析程序 geNorm 和 NormFinder 评估基因表达的稳定性。使用这些分析，RPS18 和 5.8S rRNA 无论在四个不同的品系中，其表达都是最稳定的，而 RPS18 和α-TUB 在不同的发育阶段表达最稳定。
