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Selection of reference genes for gene expression studies in Siberian Apricot (Prunus sibirica L.) Germplasm using quantitative real-time PCR.

Niu J, Zhu B, Cai J, Li P, Wang L, Dai H, Qiu L, Yu H, Ha D, Zhao H, Zhang Z, Lin S - PLoS ONE (2014)

Bottom Line: Moreover, UBC was selected as the most stably expressed gene by both algorithms in different Siberian Apricot seed samples.We also detected that a set of three genes (ACT, CYP and UBC) by geNorm as control for normalization could lead to accurate results.These obtained experimental results could make an important contribution to normalize real-time PCR data for gene expression analysis in Siberian Apricot Germplasm.

View Article: PubMed Central - PubMed

Affiliation: College of Biological Sciences and Biotechnology, College of Nature Conservation, National Engineering Laboratory for Tree Breeding, Key Laboratory of Genetics and Breeding in Forest Trees and Ornamental Plants, Ministry of Education, Beijing Forestry University, Beijing, China.

ABSTRACT
Quantitative real time reverse transcription polymerase chain reaction has been applied in a vast range of studies of gene expression analysis. However, real-time PCR data must be normalized with one or more reference genes. In this study, eleven putative consistently expressed genes (ACT, TUA, TUB, CYP, DNAj, ELFA, F-box27, RPL12, GAPDH, UBC and UBQ) in nine Siberian Apricot Germplasms (including much variability) were evaluated for their potential as references for the normalization of gene expression by NormFinder and geNorm programs. From our studies, ACT, UBC, CYP, UBQ and RPL12 as suitable for normalization were identified by geNorm, while UBC and CYP as the best pair by NormFinder. Moreover, UBC was selected as the most stably expressed gene by both algorithms in different Siberian Apricot seed samples. We also detected that a set of three genes (ACT, CYP and UBC) by geNorm as control for normalization could lead to accurate results. Furthermore, the expression levels of oleosin gene were analyzed to validate the suitability of the selected reference genes. These obtained experimental results could make an important contribution to normalize real-time PCR data for gene expression analysis in Siberian Apricot Germplasm.

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NormFinder analysis of candidate reference genes in Siberian Apricot Germplasms.
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pone-0103900-g005: NormFinder analysis of candidate reference genes in Siberian Apricot Germplasms.

Mentions: For a more robust analysis, we used the NormFinder approach, which is less affected by correlated expression compared with geNorm. A low stability value is indicative of stable expression; the NormFinder analysis provided roughly the same ranking of the candidate genes as geNorm without TUB (Figure 5). Based on our data, NormFinder identified the pair of UBC and CYP as the best one combination of reference genes in different Siberian Apricot Germplasms, and F-box27, ELFA, DNAj ranked consistently poorly.


Selection of reference genes for gene expression studies in Siberian Apricot (Prunus sibirica L.) Germplasm using quantitative real-time PCR.

Niu J, Zhu B, Cai J, Li P, Wang L, Dai H, Qiu L, Yu H, Ha D, Zhao H, Zhang Z, Lin S - PLoS ONE (2014)

NormFinder analysis of candidate reference genes in Siberian Apricot Germplasms.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4126684&req=5

pone-0103900-g005: NormFinder analysis of candidate reference genes in Siberian Apricot Germplasms.
Mentions: For a more robust analysis, we used the NormFinder approach, which is less affected by correlated expression compared with geNorm. A low stability value is indicative of stable expression; the NormFinder analysis provided roughly the same ranking of the candidate genes as geNorm without TUB (Figure 5). Based on our data, NormFinder identified the pair of UBC and CYP as the best one combination of reference genes in different Siberian Apricot Germplasms, and F-box27, ELFA, DNAj ranked consistently poorly.

Bottom Line: Moreover, UBC was selected as the most stably expressed gene by both algorithms in different Siberian Apricot seed samples.We also detected that a set of three genes (ACT, CYP and UBC) by geNorm as control for normalization could lead to accurate results.These obtained experimental results could make an important contribution to normalize real-time PCR data for gene expression analysis in Siberian Apricot Germplasm.

View Article: PubMed Central - PubMed

Affiliation: College of Biological Sciences and Biotechnology, College of Nature Conservation, National Engineering Laboratory for Tree Breeding, Key Laboratory of Genetics and Breeding in Forest Trees and Ornamental Plants, Ministry of Education, Beijing Forestry University, Beijing, China.

ABSTRACT
Quantitative real time reverse transcription polymerase chain reaction has been applied in a vast range of studies of gene expression analysis. However, real-time PCR data must be normalized with one or more reference genes. In this study, eleven putative consistently expressed genes (ACT, TUA, TUB, CYP, DNAj, ELFA, F-box27, RPL12, GAPDH, UBC and UBQ) in nine Siberian Apricot Germplasms (including much variability) were evaluated for their potential as references for the normalization of gene expression by NormFinder and geNorm programs. From our studies, ACT, UBC, CYP, UBQ and RPL12 as suitable for normalization were identified by geNorm, while UBC and CYP as the best pair by NormFinder. Moreover, UBC was selected as the most stably expressed gene by both algorithms in different Siberian Apricot seed samples. We also detected that a set of three genes (ACT, CYP and UBC) by geNorm as control for normalization could lead to accurate results. Furthermore, the expression levels of oleosin gene were analyzed to validate the suitability of the selected reference genes. These obtained experimental results could make an important contribution to normalize real-time PCR data for gene expression analysis in Siberian Apricot Germplasm.

Show MeSH