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Electronic Journal of Biotechnology
Universidad Católica de Valparaíso
ISSN: 0717-3458
Vol. 13, No. 5, 2010
Bioline Code: ej10066
Full paper language: English
Document type: Note
Document available free of charge

Electronic Journal of Biotechnology, Vol. 13, No. 5, 2010

 en Isolation of total RNA from hard bamboo tissue rich in polyphenols and polysaccharides for gene expression studies
Rai, Vineeta; Ghosh, Jayadri Sekhar & Dey, Nrisingha

Abstract

RNA isolation from hard and woody internodal bamboo ( Bambusa balcooa check for this species in other resources ) tissue is very difficult due to the presence of secondary metabolites, polysaccharides, and polyphenolics. These compounds often co-precipitate with isolated RNA and hinder downstream applications. We have developed an efficient, cost effective and reproducible RNA isolation method from hard tissue of bamboo internode. This protocol includes an additional organic solvent refinement steps to remove endogenous phenolic compounds and acidic phenol (pH 4.2) to critically stabilize RNA in extraction buffer. In addition to these, two 2M Lithium chloride washing steps were introduced to eliminate DNA and polysaccharides contamination. The RNA isolated from the present protocol was found to be superior, when compared to total RNA extracted by other available protocols. The A260/A280 absorption ratio of the isolated RNA was found ranging between 1.89-1.97. The integrity of 28S and 18S rRNA was highly satisfactory when analyzed in agarose denaturing gel. RNA was further used for RT PCR, northern hybridization, cDNA library and subtractive hybridization without any further refinement.

Keywords
acid phenol, Bambusa balcooa, fiber specific genes, internode, RNA extraction.

 
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