NDLI: An efficient RNA isolation procedure and identification of reference genes for normalization of gene expression in blueberry

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An efficient RNA isolation procedure and identification of reference genes for normalization of gene expression in blueberry

Content Provider	Springer Nature Link
Author	Vashisth, Tripti Johnson, Lisa Klima Malladi, Anish
Copyright Year	2011
Abstract	Application of transcriptomics approaches can greatly enhance our understanding of blueberry physiology. The success of transcriptomics approaches is dependent on the extraction of high-quality RNA which is complicated by the abundance of polyphenolics and polysaccharides in blueberry. Additionally, transcriptomics requires the accurate quantification of transcript abundance. Quantitative real-time polymerase chain reaction (qRT-PCR) is a robust method to determine transcript abundance. Normalization of gene expression using stably expressed reference genes is essential in qRT-PCR. An evaluation of the stability of expression of reference genes has not yet been reported in blueberry. The objectives of this study were to develop an effective procedure for extracting RNA from different organs and to evaluate potential reference genes for qRT-PCR analyses in blueberry. RNA of high quality and yield was extracted from eight and six organs of rabbiteye and southern highbush blueberry, respectively, using a modified cetyltrimethyl ammonium bromide-based method. The expression stability of 12 reference genes was evaluated. UBIQUITIN-CONJUGATING ENZYME (UBC28), RNA HELICASE-LIKE (RH8), CLATHRIN ADAPTER COMPLEXES MEDIUM SUBUNIT FAMILY PROTEIN (CACSa), and POLYUBIQUITIN (UBQ3b) were the most stably expressed genes across multiple organs in both blueberry species. Further, the expression stability of the reference genes in the branch abscission zone following treatment with fruit abscission-inducing compounds was analyzed. CACSa, RH8, and UBC28 were the most stably expressed genes in the abscission zone under abscission-inducing conditions. We suggest a preliminary evaluation of UBC28, CACSa, RH8, and UBQ3b to identify the most suitable reference genes for the experimental conditions under consideration in blueberry.
Starting Page	2167
Ending Page	2176
Page Count	10
File Format	PDF
ISSN	07217714
Journal	Plant Cell Reports
Volume Number	30
Issue Number	12
e-ISSN	1432203X
Language	English
Publisher	Springer-Verlag
Publisher Date	2011-07-15
Publisher Place	Berlin, Heidelberg
Access Restriction	One Nation One Subscription (ONOS)
Subject Keyword	Abscission CTAB RNA extraction qRT-PCR Reference genes Plant Biochemistry Cell Biology Biotechnology Plant Sciences
Content Type	Text
Resource Type	Article
Subject	Plant Science Medicine Agronomy and Crop Science

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