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Efficient protocol for isolation of functional RNA from different grape tissue rich in polyphenols and polysaccharides for gene expression studies Hemanth K.N. Vasanthaiah* Ramesh Katam Mehboob B. Sheikh Website: http://www.famu.edu/oldsite/acad/colleges/cesta/vit-faculty.htm *Corresponding author Financial support: The research was supported by Florida Grape Growers Association. Keywords: differential display RT-PCR, grape, RNA isolation, RT-PCR, subtractive hybridization, Vitis.
Extraction of RNA from plant tissue containing high levels of polyphenols and polysaccharides is tedious and difficult in grapes. Although several protocols have been published for plant RNA isolation, most have failed to yield high quality RNA in sufficient quantity from mature and diseased grape tissue. We describe a protocol for isolating intact and high quality RNA from various grape tissues as evident by high A260/A280 absorbance ratio (1.8 to 1.9) and electrophoretic profile on denaturing formaldehyde agarose gel. On an average, 205 µg RNA per g of fresh tissue were obtained using this modified protocol. RNA quality was further assessed through RT-PCR, differential display RT-PCR and subtractive hybridization, and found to be suitable for molecular studies. |