Limits...
Stable Reference Gene Selection for RT-qPCR Analysis in Nonviruliferous and Viruliferous Frankliniella occidentalis.

Yang C, Li H, Pan H, Ma Y, Zhang D, Liu Y, Zhang Z, Zheng C, Chu D - PLoS ONE (2015)

Bottom Line: To facilitate gene expression studies, normalization of genes of interest relative to stable reference genes is crucial.Additionally, two reference genes were sufficient for reliable normalization in nonviruliferous and viruliferous F. occidentalis.This work provides a foundation for investigating the molecular mechanisms of TSWV and F. occidentalis interactions.

View Article: PubMed Central - PubMed

Affiliation: Hunan Academy of Agricultural Sciences, Institute of Plant Protection, Changsha, Hunan, China.

ABSTRACT
Reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) is a reliable technique for measuring and evaluating gene expression during variable biological processes. To facilitate gene expression studies, normalization of genes of interest relative to stable reference genes is crucial. The western flower thrips Frankliniella occidentalis (Pergande) (Thysanoptera: Thripidae), the main vector of tomato spotted wilt virus (TSWV), is a destructive invasive species. In this study, the expression profiles of 11 candidate reference genes from nonviruliferous and viruliferous F. occidentalis were investigated. Five distinct algorithms, geNorm, NormFinder, BestKeeper, the ΔCt method, and RefFinder, were used to determine the performance of these genes. geNorm, NormFinder, BestKeeper, and RefFinder identified heat shock protein 70 (HSP70), heat shock protein 60 (HSP60), elongation factor 1 α, and ribosomal protein l32 (RPL32) as the most stable reference genes, and the ΔCt method identified HSP60, HSP70, RPL32, and heat shock protein 90 as the most stable reference genes. Additionally, two reference genes were sufficient for reliable normalization in nonviruliferous and viruliferous F. occidentalis. This work provides a foundation for investigating the molecular mechanisms of TSWV and F. occidentalis interactions.

No MeSH data available.


Related in: MedlinePlus

Expression profiles of 11 candidate reference genes in Frankliniella occidentalis.The expression levels of the candidate reference genes are documented by the Ct values. The median is represented by the line in the box. The interquartile rang is bordered by the upper and lower edges, which indicate the 75th and 25th percentiles, respectively.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4526564&req=5

pone.0135207.g001: Expression profiles of 11 candidate reference genes in Frankliniella occidentalis.The expression levels of the candidate reference genes are documented by the Ct values. The median is represented by the line in the box. The interquartile rang is bordered by the upper and lower edges, which indicate the 75th and 25th percentiles, respectively.

Mentions: The mean and standard deviation (SD) of Ct values were calculated for all samples (S1 Table). EF1A (SD = 0.68) had the least variable expression and this was reflected in its low SD values. By contrast, 28S (SD = 1.35) had the most variable expression, as shown by its high SD values. Additionally, 18S had the lowest Ct values (Ctavg = 10.09), suggesting that it had the highest expression level, whereas Actin was the lowest expressed gene among the candidates (Ctavg = 31.62) (Fig 1; S1 Table).


Stable Reference Gene Selection for RT-qPCR Analysis in Nonviruliferous and Viruliferous Frankliniella occidentalis.

Yang C, Li H, Pan H, Ma Y, Zhang D, Liu Y, Zhang Z, Zheng C, Chu D - PLoS ONE (2015)

Expression profiles of 11 candidate reference genes in Frankliniella occidentalis.The expression levels of the candidate reference genes are documented by the Ct values. The median is represented by the line in the box. The interquartile rang is bordered by the upper and lower edges, which indicate the 75th and 25th percentiles, respectively.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4526564&req=5

pone.0135207.g001: Expression profiles of 11 candidate reference genes in Frankliniella occidentalis.The expression levels of the candidate reference genes are documented by the Ct values. The median is represented by the line in the box. The interquartile rang is bordered by the upper and lower edges, which indicate the 75th and 25th percentiles, respectively.
Mentions: The mean and standard deviation (SD) of Ct values were calculated for all samples (S1 Table). EF1A (SD = 0.68) had the least variable expression and this was reflected in its low SD values. By contrast, 28S (SD = 1.35) had the most variable expression, as shown by its high SD values. Additionally, 18S had the lowest Ct values (Ctavg = 10.09), suggesting that it had the highest expression level, whereas Actin was the lowest expressed gene among the candidates (Ctavg = 31.62) (Fig 1; S1 Table).

Bottom Line: To facilitate gene expression studies, normalization of genes of interest relative to stable reference genes is crucial.Additionally, two reference genes were sufficient for reliable normalization in nonviruliferous and viruliferous F. occidentalis.This work provides a foundation for investigating the molecular mechanisms of TSWV and F. occidentalis interactions.

View Article: PubMed Central - PubMed

Affiliation: Hunan Academy of Agricultural Sciences, Institute of Plant Protection, Changsha, Hunan, China.

ABSTRACT
Reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) is a reliable technique for measuring and evaluating gene expression during variable biological processes. To facilitate gene expression studies, normalization of genes of interest relative to stable reference genes is crucial. The western flower thrips Frankliniella occidentalis (Pergande) (Thysanoptera: Thripidae), the main vector of tomato spotted wilt virus (TSWV), is a destructive invasive species. In this study, the expression profiles of 11 candidate reference genes from nonviruliferous and viruliferous F. occidentalis were investigated. Five distinct algorithms, geNorm, NormFinder, BestKeeper, the ΔCt method, and RefFinder, were used to determine the performance of these genes. geNorm, NormFinder, BestKeeper, and RefFinder identified heat shock protein 70 (HSP70), heat shock protein 60 (HSP60), elongation factor 1 α, and ribosomal protein l32 (RPL32) as the most stable reference genes, and the ΔCt method identified HSP60, HSP70, RPL32, and heat shock protein 90 as the most stable reference genes. Additionally, two reference genes were sufficient for reliable normalization in nonviruliferous and viruliferous F. occidentalis. This work provides a foundation for investigating the molecular mechanisms of TSWV and F. occidentalis interactions.

No MeSH data available.


Related in: MedlinePlus