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Assessment of suitable reference genes for qRT-PCR analysis in Adelphocoris suturalis
LUO Jing, MA Chao, LI Zhe, ZHU Bang-qin, ZHANG Jiang, LEI Chao-liang, JIN Shuang-xia, J. Joe Hull, CHEN Li-zhen
2018, 17 (12): 2745-2757.   DOI: 10.1016/S2095-3119(18)61926-4
Abstract258)      PDF (1312KB)(318)      
Quantitative reverse transcription polymerase chain reaction (qRT-PCR) is the most commonly-used tool for measurement of gene expression, but its accuracy and reliability depend on appropriate data normalization with the use of one or more stable reference genes.  Adelphocoris suturalis is one of the most destructive pests of cotton, but until recently knowledge of its underlying molecular physiology had been hindered by a lack of molecular resources.  To facilitate research on this pest, we evaluated 12 common housekeeping genes studied in insects (GAPDH, ACT, βACT, TBP, SDH, βTUB, EF1γ, EF1α, EF1δ, RPL32, RPS15, and RPL27) for their expression stability in A. suturalis when subjected to various experimental treatments, including three biotic (developmental stage and sex, tissue type, and metathoracic scent gland for varying developmental stages and sexes) and one abiotic (RNA interference injection) conditions.  Four dedicated algorithms (ΔCt method, geNorm, BestKeeper and NormFinder) were used to analyze gene expression stability.  In addition, RefFinder provided an overall ranking of the stability/suitability of these candidates.  This study is the first to provide a comprehensive list of suitable reference genes for gene expression analyses in A. suturalis, which can serve to facilitate transcript expression study of related biological processes in this and related species.
 
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DNA methylation patterns of banana leaves in response to Fusarium oxysporum f. sp. cubense tropical race 4
LUO Jing-yao, PAN Xiao-lei, PENG Tie-cheng, CHEN Yun-yun, ZHAO Hui, MU Lei, PENG Yun, HE Rui, TANG Hua
2016, 15 (12): 2736-2744.   DOI: 10.1016/S2095-3119(16)61495-8
Abstract1275)      PDF in ScienceDirect      
    Fusarium wilt of banana, which is caused by Fusarium oxysporum f. sp. cubense tropical race 4 (Foc TR4), is a serious soil-borne fungal disease. Now, the epigenetic molecular pathogenic basis is elusive. In this study, with methylation-sensitive amplification polymorphism (MSAP) technique, DNA methylation was compared between the leaves inoculated with Foc TR4 and the mock-inoculated leaves at different pathogenic stages. With 25 pairs of primers, 1 144 and 1 255 fragments were amplified from the infected and mock-inoculated leaves, respectively. DNA methylation was both changed and the average methylated CCGG sequences were 34.81 and 29.26% for the infected and the mock-inoculated leaves. And DNA hypermethylation and hypomethylation were induced by pathogen infection during all pathogenic stages. Further, 69 polymorphic fragments were sequenced and 29 of them showed sequence similarity to genes with known functions. And RT-PCR results of four genes indicated that their expression patterns were consistent with their methylation patterns. Our results suggest that DNA methylation plays important roles in pathogenic response to Foc TR4 for banana.
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