Cy. Wan et Ta. Wilkins, A MODIFIED HOT BORATE METHOD SIGNIFICANTLY ENHANCES THE YIELD OF HIGH-QUALITY RNA FROM COTTON (GOSSYPIUM-HIRSUTUM L), Analytical biochemistry, 223(1), 1994, pp. 7-12
The isolation of biologically active RNA from cotton (Gossypium hirsut
um L.) is difficult due to interference by high levels of endogenous p
henolics, polysaccharides, and secondary metabolites. A modified hot b
erate procedure was developed to combat these cellular constituents du
ring tissue homogenization, resulting in the quantitative recovery of
RNA suitable for hybridization analysis, in vitro translation, and cDN
A synthesis. The efficacy of several hot berate buffer adjuvants for t
he qualitative and quantitative recovery of leaf RNA was monitored by
absorbance spectra, gel electrophoresis, protein, and cDNA synthesis.
Of the buffer adjuvants evaluated, polyvinylpyrrolidone-40 (PVP-40) ex
hibited the single, most significant impact on the yield and quality o
f RNA isolated from cotton leaves, although inclusion of deoxycholate
and/or Nonident-40 (NP-40) further enhanced the quality of the RNA. Th
e unsurpassed qualitative and quantitative recovery of total RNA from
cotton by hot berate buffer at alkaline pH, supplemented with PVP-40,
deoxycholate, and/or NP-40 had also proven satisfactory for other reca
lcitrant plant species as well as for especially difficult tissue type
s.