Identification and validation of reference genes for normalization of gene expression analysis using qRT-PCR in Helicoverpa armigera (Lepidoptera: Noctuidae)

Zhang, Songdou; An, Shiheng; Li, Zhen; Wu, Fengming; Yang, Qingliang; Liu, Yichen; Cao, Jinjun; Zhang, Huaijiang; Zhang, Qingwen; Liu, Xiaoxia

doi:10.1016/j.gene.2014.11.038

Cited by 119 publications

(119 citation statements)

References 53 publications

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“…However, GAPDH was not stable in Petunia x hybrida during leaf and flower development (Mallona et al 2010). EF1α was also recommended as one of the most stable reference gene to validate gene expression data in this study, the similar results were obtained in other researches (Hornáková et al 2010;Zhang et al 2015). The suitability rankings of the reference genes were different with the different programs as the stability of the expression of eight candidate reference genes was evaluated via five commonly used programs (RefFinder, geNorm, NormFinder, BestKeeper, and ΔCt method) for data generated in different experimental conditions.…”

Section: Discussionsupporting

confidence: 79%

Selection and evaluation of potential reference genes for gene expression analysis in Avena fatua

Liu¹,

Li²,

Lu³

et al. 2019

Plant Prot. Sci.

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Citation: Liu J., Li P., Lu L., Xie L., Chen X., Zhang B. (2019): Selection and evaluation of potential reference genes for gene expression analysis in Avena fatua. Plant Protect. Sci., 55: 61-71.Abstract: Eight commonly used candidate reference genes, 18S ribosomal RNA (rRNA) (18S), 28S rRNA (28S), actin (ACT), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), elongation factor 1 alpha (EF1α), ribosomal protein L7 (RPL7), Alpha-tubulin (α-TUB), and TATA box binding protein-associated factor (TBP), were evaluated under various experimental conditions to assess their suitability in different developmental stages, tissues and herbicide treatments in Avena fatua. The results indicated the most suitable reference genes for the different experimental conditions. For developmental stages, 28S and EF1α were the optimal reference genes, both EF1α and 28S were suitable for experiments of different tissues, whereas for herbicide treatments, GAPDH and ACT were suitable for normalizations of expression data. In addition, GAPDH and EF1α were the suitable reference genes.

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Section: Discussionsupporting

confidence: 79%

Selection and evaluation of potential reference genes for gene expression analysis in Avena fatua

Liu¹,

Li²,

Lu³

et al. 2019

Plant Prot. Sci.

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“…Two ribosomal protein genes ( RPS15 and RPS13 ) also showed stable expressions among five Gynaephora populations, as observed in Helicoverpa armigera under different developmental stages and tissues (Zhang et al., 2014). RPS15 had been demonstrated to be a RG in humans (Kitagawa et al., 1991; Shiga, Yamamoto, & Okamoto, 1990) and is a suitable RG in many cases in mammals (Bionaz & Loor, 2007; Kumar et al., 2012).…”

Section: Discussionmentioning

confidence: 99%

Selection of reference genes for qRT‐PCR and expression analysis of high‐altitude‐related genes in grassland caterpillars (Lepidoptera: Erebidae: Gynaephora) along an altitude gradient

Zhang

Wang

et al. 2017

Ecology and Evolution

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Changes in gene expression patterns can reflect the adaptation of organisms to divergent environments. Quantitative real‐time PCR (qRT‐PCR) is an important tool for ecological adaptation studies at the gene expression level. The quality of the results of qRT‐PCR analysis largely depends on the availability of reliable reference genes (RGs). To date, reliable RGs have not been determined for adaptive evolution studies in insects using a standard approach. Here, we evaluated the reliability of 17 candidate RGs for five Gynaephora populations inhabiting various altitudes of the Tibetan Plateau (TP) using four independent (geNorm, NormFinder, BestKeeper, and the deltaCt method) and one comprehensive (RefFinder) algorithms. Our results showed that EF1‐α, RPS15, and RPS13 were the top three most suitable RGs, and a combination of these three RGs was the most optimal for normalization. Conversely, RPS2,ACT, and RPL27 were the most unstable RGs. The expression profiles of two target genes (HSP70 and HSP90) were used to confirm the reliability of the chosen RGs. Additionally, the expression patterns of four other genes (GPI,HIF1A,HSP20, and USP) associated with adaptation to extreme environments were assessed to explore the adaptive mechanisms of TP Gynaephora species to divergent environments. Each of these six target genes showed discrepant expression patterns among the five populations, suggesting that the observed expression differences may be associated with the local adaptation of Gynaephora to divergent altitudinal environments. This study is a useful resource for studying the adaptive evolution of TP Gynaephora to divergent environments using qRT‐PCR, and it also acts as a guide for selecting suitable RGs for ecological and evolutionary studies in insects.

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“…With more and more genomes and transcriptomes available for arthropods, there are at least 45 arthropod species that have been investigated for reference gene identification and selection under diverse experimental conditions. These arthropods consist of various agricultural and urban pests 39-50, beneficial predators 51, 52, pollinators 53-56, and other economically important insects such as silkworm 57 (also see Table S4). Most of these studies compared gene expression across several biotic or abiotic variables, such as developmental stage, tissues type, pesticide exposure, and temperature stress.…”

Section: Discussionmentioning

confidence: 99%

Selection of Reference Genes for Expression Studies of Xenobiotic Adaptation in Tetranychus urticae

et al. 2016

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Quantitative real-time PCR (qRT-PCR) is an extensively used, high-throughput method to analyze transcriptional expression of genes of interest. An appropriate normalization strategy with reliable reference genes is required for calculating gene expression across diverse experimental conditions. In this study, we aim to identify the most stable reference genes for expression studies of xenobiotic adaptation in Tetranychus urticae, an extremely polyphagous herbivore causing significant yield reduction of agriculture. We chose eight commonly used housekeeping genes as candidates. The qRT-PCR expression data for these genes were evaluated from seven populations: a susceptible and three acaricide resistant populations feeding on lima beans, and three other susceptible populations which had been shifted host from lima beans to three other plant species. The stability of the candidate reference genes was then assessed using four different algorithms (comparative ΔCt method, geNorm, NormFinder, and BestKeeper). Additionally, we used an online web-based tool (RefFinder) to assign an overall final rank for each candidate gene. Our study found that CycA and Rp49 are best for investigating gene expression in acaricide susceptible and resistant populations. GAPDH, Rp49, and Rpl18 are best for host plant shift studies. And GAPDH and Rp49 were the most stable reference genes when investigating gene expression under changes in both experimental conditions. These results will facilitate research in revealing molecular mechanisms underlying the xenobiotic adaptation of this notorious agricultural pest.

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Identification and validation of reference genes for normalization of gene expression analysis using qRT-PCR in Helicoverpa armigera (Lepidoptera: Noctuidae)

Cited by 119 publications

References 53 publications

Selection and evaluation of potential reference genes for gene expression analysis in Avena fatua

Selection and evaluation of potential reference genes for gene expression analysis in Avena fatua

Selection of reference genes for qRT‐PCR and expression analysis of high‐altitude‐related genes in grassland caterpillars (Lepidoptera: Erebidae: Gynaephora) along an altitude gradient

Selection of Reference Genes for Expression Studies of Xenobiotic Adaptation in Tetranychus urticae

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