Selection of Endogenous Reference Genes for Gene Expression Analysis in the Mediterranean Species of the &lt;I&gt;Bemisia tabaci&lt;/I&gt; (Hemiptera: Aleyrodidae) Complex

Yan, Su; He, Wenbo; Wang, Jia; Li, Junmin; Liu, Shu‐Sheng; Wang, Xiaowei

doi:10.1603/ec12459

Cited by 28 publications

(26 citation statements)

References 58 publications

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“…tabaci [18, 21]. While, primer sequences of β-tub were designed from the conserved regions of related sequences available in the transcriptome of B .…”

Section: Resultsmentioning

confidence: 99%

“…In order to overcome the limitation of this method, a robust normalization strategy is utilized by using reference genes, whose expression levels should remain stable independent of tissue types, developmental stages and experimental conditions [16, 17]. Nevertheless, previous experiments evidently showed that the expression of universal reference genes in arthropods could be variable, particularly in different tissues, developmental stages and experimental conditions such as length of exposure to distinct abiotic factors [18–21]. Moreover, normalisation against a single reference gene has been found to be insufficient without the selection of multiple reference genes [15, 22, 23].…”

Section: Introductionmentioning

confidence: 99%

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Selection and validation of reference genes for qRT-PCR analysis during biological invasions: The thermal adaptability of Bemisia tabaci MED

Dai

Liu

et al. 2017

PLoS ONE

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The Bemisia tabaci Mediterranean (MED) cryptic species has been rapidly invading to most parts of the world owing to its strong ecological adaptability, which is considered as a model insect for stress tolerance studies under rapidly changing environments. Selection of a suitable reference gene for quantitative stress-responsive gene expression analysis based on qRT-PCR is critical for elaborating the molecular mechanisms of thermotolerance. To obtain accurate and reliable normalization data in MED, eight candidate reference genes (β-act, GAPDH, β-tub, EF1-α, GST, 18S, RPL13A and α-tub) were examined under various thermal stresses for varied time periods by using geNorm, NormFinder and BestKeeper algorithms, respectively. Our results revealed that β-tub and EF1-α were the best reference genes across all sample sets. On the other hand, 18S and GADPH showed the least stability for all the samples studied. β-act was proved to be highly stable only in case of short-term thermal stresses. To our knowledge this was the first comprehensive report on validation of reference genes under varying temperature stresses in MED. The study could expedite particular discovery of thermotolerance genes in MED. Further, the present results can form the basis of further research on suitable reference genes in this invasive insect and will facilitate transcript profiling in other invasive insects.

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“…tabaci [18, 21]. While, primer sequences of β-tub were designed from the conserved regions of related sequences available in the transcriptome of B .…”

Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Selection and validation of reference genes for qRT-PCR analysis during biological invasions: The thermal adaptability of Bemisia tabaci MED

Dai

Liu

et al. 2017

PLoS ONE

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“…Similarly, Actin was never ranked as a suitable reference gene across different developmental stages in T. urticae 62, or across different developmental stages and abiotic stresses in other mite species T. cinnabarinus 63 and Panonychus citri 64. Actin also showed extreme variation in expression throughout developmental stages of the silkworm Bombyx mori 57, and in different physiological conditions in the whitefly Bemisia tabaci 65. In contrast, RP49 , which has also been used in previous studies, was ranked as the second most stable reference gene across all experimental conditions in our study (Table 3).…”

Section: Discussionmentioning

confidence: 99%

“…In the same species, the best reference gene sets can vary significantly under different biotic and abiotic conditions 46, 65. Even under a single experimental condition (e.g.…”

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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“…The relative expression levels of each gene in gut and body were normalized using the threshold cycle (Ct) values that were obtained from reactions run on the same plate. TATA Box Binding Protein-Associated Factor (TAF) was chosen as the endogenous reference gene in qPCR analysis with the 2 -△△Ct method [60, 61]. To confirm the results of RPKM analyses of orthologous gene expression between MEAM1 and MED gut, the levels of expression of 20 selected genes (10 MEAM1 up-regulated genes and 10 MEAM1 down-regulated genes) were measured.…”

Section: Methodsmentioning

confidence: 99%

Transcriptomic analyses reveal the adaptive features and biological differences of guts from two invasive whitefly species

Yan

Zhao

et al. 2014

BMC Genomics

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BackgroundThe gut of phloem feeding insects is critical for nutrition uptake and xenobiotics degradation. However, partly due to its tiny size, genomic information for the gut of phloem feeding insects is limited.ResultsIn this study, the gut transcriptomes of two species of invasive whiteflies in the Bemisia tabaci complex, Middle East Asia Minor 1 (MEAM1) and Mediterranean (MED), were analyzed using the Illumina sequencing. A total of 12,879 MEAM1 transcripts and 11,246 MED transcripts were annotated with a significant Blastx hit. In addition, 7,000 and 5,771 gut specific genes were respectively identified for MEAM1 and MED. Functional analyses on these gut specific genes demonstrated the important roles of gut in metabolism of insecticides and secondary plant chemicals. To reveal the molecular difference between guts of MEAM1 and MED, a comparison between gut transcriptomes of the two species was conducted and 3,910 pairs of orthologous genes were identified. Based on the ratio of nonsynonymous and synonymous substitutions, 15 genes were found evolving under positive selection. Many of those genes are predicted to be involved in metabolism and insecticide resistance. Furthermore, many genes related to detoxification were expressed at an elevated level in the gut of MED compared to MEAM1, which might be responsible for the MED’s higher resistance to insecticides and environmental stresses.ConclusionThe sequencing of MED and MEAM1 gut transcriptomes and extensive comparisons of MEAM1 and MED gut transcripts provide substantial sequence information for revealing the role of gut in whiteflies.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-370) contains supplementary material, which is available to authorized users.

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Selection of Endogenous Reference Genes for Gene Expression Analysis in the Mediterranean Species of the <I>Bemisia tabaci</I> (Hemiptera: Aleyrodidae) Complex

Cited by 28 publications

References 58 publications

Selection and validation of reference genes for qRT-PCR analysis during biological invasions: The thermal adaptability of Bemisia tabaci MED

Selection and validation of reference genes for qRT-PCR analysis during biological invasions: The thermal adaptability of Bemisia tabaci MED

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

Transcriptomic analyses reveal the adaptive features and biological differences of guts from two invasive whitefly species

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