Livramento Kalynka G do, Freitas Natália C, Máximo Wesley P F, Zanetti Ronald, Paiva Luciano V
Central Laboratory of Molecular Biology, Federal University of Lavras (UFLA), Lavras, MG 37200-000, Brazil.
Entomology Department, Federal University of Lavras (UFLA), Lavras, MG 37200-000, Brazil.
Insects. 2018 Feb 8;9(1):18. doi: 10.3390/insects9010018.
Although several ant species are important targets for the development of molecular control strategies, only a few studies focus on identifying and validating reference genes for quantitative reverse transcription polymerase chain reaction (RT-qPCR) data normalization. We provide here an extensive study to identify and validate suitable reference genes for gene expression analysis in the ant , a threatening agricultural pest in South America. The optimal number of reference genes varies according to each sample and the result generated by RefFinder differed about which is the most suitable reference gene. Results suggest that the , and genes were the best reference genes in the sample pool according to stability values. The gene expression pattern was stable in all evaluated sample set. In contrast, when using less stable reference genes for normalization a large variability in gene expression was recorded. There is no universal reference gene suitable for all conditions under analysis, since these genes can also participate in different cellular functions, thus requiring a systematic validation of possible reference genes for each specific condition. The choice of reference genes on gene normalization confirmed that unstable reference genes might drastically change the expression profile analysis of target candidate genes.
虽然几种蚂蚁物种是分子控制策略开发的重要目标，但只有少数研究专注于鉴定和验证用于定量逆转录聚合酶链反应（RT-qPCR）数据标准化的参考基因。我们在此进行了一项广泛的研究，以鉴定和验证适合在红火蚁（一种南美洲具有威胁性的农业害虫）中进行基因表达分析的参考基因。参考基因的最佳数量因每个样本而异，并且RefFinder生成的结果在哪个是最合适的参考基因方面存在差异。结果表明，根据稳定性值，actin、EF1α和β-tubulin基因是样本库中最佳的参考基因。在所有评估的样本集中，EF1α基因表达模式稳定。相反，当使用稳定性较差的参考基因进行标准化时，记录到红火蚁基因表达存在很大变异性。由于这些基因也可参与不同的细胞功能，因此不存在适用于所有分析条件的通用参考基因，这就需要针对每种特定条件对可能的参考基因进行系统验证。在红火蚁基因标准化中参考基因的选择证实，不稳定的参考基因可能会极大地改变目标候选基因的表达谱分析。
