Arbitrary Multi-gene Reference for Normalization of Real-Time PCR Gene Expression Data

Tai, Helen H.; Conn, Gregory; Davidson, Campbell G.; Platt, H. W.

doi:10.1007/s11105-009-0089-0

Cited by 17 publications

(11 citation statements)

References 24 publications

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“…PCR exponentially amplifies a small starting quantity of cDNA to a large amount of DNA product; in fact, even seemingly insignificant technical variability before amplification can lead to considerable errors in the end product, although this is less of a concern in RT-qPCR than in conventional RT-PCR. To overcome this limitation, researchers typically normalized the mRNA expression of their genes of interest to the expression of a constitutively expressed reference gene (Udvardi et al 2008), and new reference genes are being considered, including synthetic templates and random oligoprimers (Phillips et al 2009;Tai et al 2009). …”

Section: Introductionmentioning

confidence: 99%

Reference Gene Selection for Real-Time Quantitative Polymerase Chain Reaction of mRNA Transcript Levels in Chinese Cabbage (Brassica rapa L. ssp. pekinensis)

et al. 2010

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The Chinese cabbage is a crop belonging to the family Cruciferae; very few studies have focused on the analysis of gene expression in this economically important crop. In this study, we used real-time quantitative polymerase chain reaction to identify the control genes that are the most stably expressed in a given set of tissues and under conditions of drought stress and downy mildew infection. We characterized the transcript stability of nine candidate reference genes, namely, those encoding glyceraldehyde-3-phosphate dehydrogenase (GAPDH), ubiquitin (Ubc), elongation-factor-1-α (EF-1-α), adenine phosphoribosyltransferase (Apr), clathrin, cyclophilin (Cyp), tubulin (Tub), Actin, and 18s rRNA, using the geNorm and Normfinder software programs. The results of a geNorm analysis indicated that EF-1-α and Apr were the most suitable reference genes among the given set of tissues and that GAPDH and Ubc were the most stable genes under conditions of drought stress and downy mildew infection. Furthermore, the results of an analysis performed using the Normfinder software program indicated EF-1-α to be the best normalization factor in the given set of tissues and under conditions of downy mildew infection and GAPDH to be the best factor under drought stress conditions. The results of the geNorm analysis also helped determine the minimum number of genes required to calculate a reliable normalization factor. A study on the variability of expression of PSY expression showed that the relative quantification of this gene varied depending on the internal control and the number of internal controls used, thus highlighting the importance of the choice of internal controls in such experiments.

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Section: Introductionmentioning

confidence: 99%

Reference Gene Selection for Real-Time Quantitative Polymerase Chain Reaction of mRNA Transcript Levels in Chinese Cabbage (Brassica rapa L. ssp. pekinensis)

et al. 2010

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“…Quantitative real-time polymerase chain reaction, a precise method to measure changes in the gene transcription level, has been widely used in recent years (Remans et al 2008;Phillips et al 2009;Tai et al 2009;Qi et al 2010). The upregulated expression pattern of the MxbHLH01 gene was similar to that of AtFRU accumulating to high levels under iron-deficient conditions in roots (Jakoby et al 2004), but this pattern was different from that of LeFER, which was reported to be independent of the iron supply (Ling et al 2002).…”

Section: Discussionmentioning

confidence: 99%

Isolation and Characterization of the Iron-Regulated MxbHLH01 Gene in Malus xiaojinensis

Wang

Chen

et al. 2011

Plant Mol Biol Rep

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“…The 27 genes included 5 housekeeping genes and 22 experimental genes ( Table 1). The five housekeeping genes were adenine phosphoribosyl transferase (aprt), tubulin, cyclophilin (Nicot et al 2005;Tai et al 2009), elongation factor 1-α(ef1α) (Nakane et al 2003) and COX1 (Quiñones et al 1995;Li et al 1996). The test genes are described in Table 1.…”

Section: Rna Extraction and Quantification Of Gene Expressionmentioning

confidence: 99%

Differential gene expression as an indicator of nitrogen sufficiency in field-grown potato plants

et al. 2011

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Use of an in-season measure of crop N sufficiency to guide fertilizer management is one approach to match the supply of N to the crop N demand. This study examined use of gene expression in leaf tissue of field-grown potatoes for use in assessment of potato N sufficiency. Potato cultivar 'Shepody' was grown with six fertilizer N rates (0-250 kg N ha -1 ). Leaf disks were collected weekly for quantification of the expression of N uptake/transport, N assimilation, and amino acid metabolism genes in leaf tissue by nCounter. Many of the genes evaluated were responsive to crop N supply, but the response varied widely among sampling dates. The exception was an ammonium transporter gene (AT1) which was highly expressed, was relatively consistent across sampling dates, was closely related to root zone soil nitrate concentration across N rates and sampling dates, and was highly negatively correlated with total tuber yield. The level of expression of AT1 in leaf tissue was as good as or better than conventional chemical or optical measures of potato N sufficiency in the current study.

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Arbitrary Multi-gene Reference for Normalization of Real-Time PCR Gene Expression Data

Cited by 17 publications

References 24 publications

Reference Gene Selection for Real-Time Quantitative Polymerase Chain Reaction of mRNA Transcript Levels in Chinese Cabbage (Brassica rapa L. ssp. pekinensis)

Reference Gene Selection for Real-Time Quantitative Polymerase Chain Reaction of mRNA Transcript Levels in Chinese Cabbage (Brassica rapa L. ssp. pekinensis)

Isolation and Characterization of the Iron-Regulated MxbHLH01 Gene in Malus xiaojinensis

Differential gene expression as an indicator of nitrogen sufficiency in field-grown potato plants

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