Selection and Evaluation of Potential Reference Genes for Gene Expression Analysis in the Brown Planthopper, Nilaparvata lugens Hemiptera: Delphacidae Using Reverse-Transcription Quantitative PCRReportar como inadecuado




Selection and Evaluation of Potential Reference Genes for Gene Expression Analysis in the Brown Planthopper, Nilaparvata lugens Hemiptera: Delphacidae Using Reverse-Transcription Quantitative PCR - Descarga este documento en PDF. Documentación en PDF para descargar gratis. Disponible también para leer online.

The brown planthopper BPH, Nilaparvata lugens Hemiptera, Delphacidae, is one of the most important rice pests. Abundant genetic studies on BPH have been conducted using reverse-transcription quantitative real-time PCR qRT-PCR. Using qRT-PCR, the expression levels of target genes are calculated on the basis of endogenous controls. These genes need to be appropriately selected by experimentally assessing whether they are stably expressed under different conditions. However, such studies on potential reference genes in N. lugens are lacking. In this paper, we presented a systematic exploration of eight candidate reference genes in N. lugens, namely, actin 1 ACT, muscle actin MACT, ribosomal protein S11 RPS11, ribosomal protein S15e RPS15, alpha 2-tubulin TUB, elongation factor 1 delta EF, 18S ribosomal RNA 18S, and arginine kinase AK and used four alternative methods BestKeeper, geNorm, NormFinder, and the delta Ct method to evaluate the suitability of these genes as endogenous controls. We examined their expression levels among different experimental factors developmental stage, body part, geographic population, temperature variation, pesticide exposure, diet change, and starvation following the MIQE Minimum Information for publication of Quantitative real time PCR Experiments guidelines. Based on the results of RefFinder, which integrates four currently available major software programs to compare and rank the tested candidate reference genes, RPS15, RPS11, and TUB were found to be the most suitable reference genes in different developmental stages, body parts, and geographic populations, respectively. RPS15 was the most suitable gene under different temperature and diet conditions, while RPS11 was the most suitable gene under different pesticide exposure and starvation conditions. This work sheds light on establishing a standardized qRT-PCR procedure in N. lugens, and serves as a starting point for screening for reference genes for expression studies of related insects.



Autor: Miao Yuan , Yanhui Lu , Xun Zhu, Hu Wan, Muhammad Shakeel, Sha Zhan, Byung-Rae Jin, Jianhong Li

Fuente: http://plos.srce.hr/



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