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ARS Home » Pacific West Area » Aberdeen, Idaho » Small Grains and Potato Germplasm Research » Research » Publications at this Location » Publication #323224

Research Project: Genetic Improvement of Barley and Oats for Enhanced Quality and Biotic Stress Resistance

Location: Small Grains and Potato Germplasm Research

Title: Extraction of high-quality RNA from germinating barley (Hordeum vulgare L.) seeds containing high levels of starch.

Author
item Rashid, Abdur
item Gines, Michael - Brigham Young University
item Baldwin, Thomas
item Esvelt Klos, Kathy
item Bregitzer, Phil

Submitted to: Proceedings International Barley Genetics Symposium
Publication Type: Abstract Only
Publication Acceptance Date: 6/26/2016
Publication Date: 6/26/2016
Citation: Rashid, A., Gines, M., Baldwin, T.T., Esvelt Klos, K.L., Bregitzer, P.P. 2016. Extraction of high-quality RNA from germinating barley (Hordeum vulgare L.) seeds containing high levels of starch.. Proceedings International Barley Genetics Symposium. p.155.

Interpretive Summary:

Technical Abstract: Comparative evaluation of gene expression levels can lead to improved understanding of the gene networks underlying traits of economic importance. Extraction of high-quality RNA from germinating barley seeds that contain high levels of starch is of vital importance for analysing the expression of candidate genes of malting quality traits through techniques such as quantitative real-time polymerase chain reaction (qRT-PCR). Most of the existing RNA extraction protocols published in the literature for seed tissues fail to yield high quality RNA in barley because elevated levels of starch contaminate RNA by co-precipitation. This causes sample solidification and hinders RNA dissolution in nuclease-free water, reducing both yield and quality. Here we describe an extraction protocol for high quality RNA from germinating barley seeds based on Li and Trick (2005), but with substantial modifications. RNA extracted with this protocol is of excellent quality and could be used for expression analysis of genes in germinating seeds of cereal crops, including barley, by qRT-PCR.