Selection of reference genes for tissue/organ samples of adults of Eucryptorrhynchus scrobiculatus

Gao, Peng; Wang, Jingzhen; Wen, Junbao

doi:10.1371/journal.pone.0228308

Cited by 12 publications

(12 citation statements)

References 52 publications

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“…This is mainly related to the variability of reference genes in different environments and species. In addition, in the studies of reference gene stability, these commonly used reference genes are often not the most optimal choice (Chapman and Waldenström, 2015;Adeyinka et al, 2019;Gao et al, 2020), and this study has the same findings. In the qRT-PCR research on ginger, common reference genes (TUB, actin, EF1α, etc.)…”

Section: Discussionmentioning

confidence: 51%

Identification of Ginger (Zingiber officinale Roscoe) Reference Genes for Gene Expression Analysis

Liu

et al. 2020

Front. Genet.

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Quantitative real-time PCR (qRT-PCR) is widely used in the detection of gene expression level. However, there is no suitable ginger reference gene for qPCR analysis. Therefore, it is the primary task to select and validate the appropriate ginger reference gene to normalize the expression of target genes. In this study, 14 candidate reference genes were selected and analyzed in different tissues (leaf, and rhizome), different development stages, different varieties, and abiotic stress (ABA and salt stress). Expression stability was calculated using geNorm and NormFinder, Bestkeeper, and RefFinder. For abiotic stress and total conditions, 28S and COX were identified as the most stable genes. In addition, RPII was the most stable in the different development stages and different varieties. TEF2 and RPL2 were the least stably expressed in the tissue and all the conditions. In order to verify the feasibility of these genes as reference genes, we used the most stable and least stable reference genes to normalize the expression levels of ZoSPS genes under different conditions. This work can provide theoretical support for future research on ginger gene expression.

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Section: Discussionmentioning

confidence: 51%

Identification of Ginger (Zingiber officinale Roscoe) Reference Genes for Gene Expression Analysis

Liu

et al. 2020

Front. Genet.

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“…qPCR is an efficient and convenient method for gene expression analysis, and using an appropriate reference gene or genes is the basic requirement to achieve qPCR reliability. To date, the selection of reliable reference genes under different conditions has been performed in many insect and mite species [ 8 ] such as locusts [ 20 , 21 ], pea aphids [ 22 ], psocids [ 23 ], honey bees [ 24 ], wasps [ 25 ], cotton bollworms [ 26 ], flies [ 27 , 28 ], silkworms [ 29 ], moths [ 30 ], beetles [ 31 , 32 , 33 ] and spider mites [ 34 , 35 , 36 ], but not the firebrat T. domestica , a representative basal insect, which plays a key role in understanding the origin of insect metamorphosis. To develop functional genomic studies in this insect, it is essential to select optimal reference genes and test the efficacy of RNAi.…”

Section: Discussionmentioning

confidence: 99%

“…Actin , a traditional housekeeping gene , has been regarded as a stable reference gene across insect species in various experimental settings [ 20 , 21 , 24 , 37 ] but not in beetles [ 31 , 32 , 33 ]. Elongation factor 1 alpha ( EF-1α ), an evolutionarily conserved GTPase, was evaluated as the stable gene in Orthoptera [ 21 ], Drosophila [ 37 ], and Hymenoptera [ 38 ].…”

Section: Discussionmentioning

confidence: 99%

Selection of Reference Genes for Normalization of Gene Expression in Thermobia domestica (Insecta: Zygentoma: Lepismatidae)

Bai

Zeng

et al. 2020

Genes

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Zygentoma occupies a key evolutionary position for understanding the evolution of insect metamorphosis but has received little attention in terms of genetic analysis. To develop functional genomic studies in this insect, we evaluated five candidate internal reference genes for quantitative RT-PCR (qPCR) studies from Thermobia domestica, a representative species of Zygentoma, including Actin 5C (Actin5C), Elongation factor-1 alpha (EF1A), Ribosome protein S26 (RPS26), Ribosome protein L32 (RPL32), and Superoxide dismutase 2 (SOD2), at different developmental stages, in various body parts, and under dsRNA microinjection and starvation stresses, using four algorithms (delta Ct, geNorm, NormFinder and BestKeeper) and a comparative algorithm (RefFinder). Specific suitable reference genes were recommended across specific experimental conditions, and the combination of RPS26 and RPL32 was appropriate for all tested samples. Employing our selected reference gene combination, we investigated the gene expression pattern of Myoglianin (Myo), a crucial gene-regulating insect metamorphosis, in ametabolous T. domestica, and demonstrated the efficiency of RNA interference (RNAi) in firebrat nymphs. This study provides a basis for reliable quantitative studies of genes and greatly benefits evolutionary and functional genomics studies in Zygentoma.

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“…Since its publication in 2018, this article by Shakeel and colleagues on insects has been cited 22 times by articles in PubMed. Fourteen of these citing articles are those evaluating candidate reference genes [ 166 , 167 , 168 , 169 , 170 , 171 , 172 , 173 , 174 , 175 , 176 , 177 , 178 , 179 ]. We hope that our critical review will similarly stimulate future research on experimentally validating reference genes for gene expression studies in Aspergillus (and in fungi in general) using RT-qPCR.…”

Section: Concluding Remarks and Recommendationsmentioning

confidence: 99%

Current Practices for Reference Gene Selection in RT-qPCR of Aspergillus: Outlook and Recommendations for the Future

Archer

2021

Genes

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Aspergillus is a genus of filamentous fungi with vast geographic and ecological distributions. Species within this genus are clinically, agriculturally and biotechnologically relevant, leading to increasing interest in elucidating gene expression dynamics of key metabolic and physiological processes. Reverse-transcription quantitative Polymerase Chain Reaction (RT-qPCR) is a sensitive and specific method of quantifying gene expression. A crucial step for comparing RT-qPCR results between strains and experimental conditions is normalisation to experimentally validated reference gene(s). In this review, we provide a critical analysis of current reference gene selection and validation practices for RT-qPCR gene expression analyses of Aspergillus. Of 90 primary research articles obtained through our PubMed query, 17 experimentally validated the reference gene(s) used. Twenty reference genes were used across the 90 studies, with beta-tubulin being the most used reference gene, followed by actin, 18S rRNA and glyceraldehyde 3-phosphate dehydrogenase. Sixteen of the 90 studies used multiple reference genes for normalisation. Failing to experimentally validate the stability of reference genes can lead to conflicting results, as was the case for four studies. Overall, our review highlights the need to experimentally validate reference genes in RT-qPCR studies of Aspergillus.

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Selection of reference genes for tissue/organ samples of adults of Eucryptorrhynchus scrobiculatus

Cited by 12 publications

References 52 publications

Identification of Ginger (Zingiber officinale Roscoe) Reference Genes for Gene Expression Analysis

Identification of Ginger (Zingiber officinale Roscoe) Reference Genes for Gene Expression Analysis

Selection of Reference Genes for Normalization of Gene Expression in Thermobia domestica (Insecta: Zygentoma: Lepismatidae)

Current Practices for Reference Gene Selection in RT-qPCR of Aspergillus: Outlook and Recommendations for the Future

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