African Journal of
Biotechnology

  • Abbreviation: Afr. J. Biotechnol.
  • Language: English
  • ISSN: 1684-5315
  • DOI: 10.5897/AJB
  • Start Year: 2002
  • Published Articles: 12496

Full Length Research Paper

A mini-scale hot borate method for the isolation of total RNA from a large number of cotton tissue samples

Mingxiong Pang1, J. McD. Stewart2  and Jinfa Zhang1*
1Department of Plant and Environmental Sciences, New Mexico State University, Las Cruces, NM 88003, New Mexico, USA. 2Department of Crop, Soil and Environmental Sciences, University of Arkansas, Fayetteville, AR 72701, USA.
Email: [email protected]

  •  Accepted: 19 September 2011
  •  Published: 02 November 2011

Abstract

Genetic genomics in cotton requires isolating RNA from a large number of plant tissue samples, in a mapping population. However, traditional methods for the extraction of RNA from cotton tissues is time consuming and not suitable for processing many samples at the same time. Here, we present a mini-scale protocol for quick isolation of total RNA from cotton ovaries with high yield and quality suitable for gene expression studies. By modifying and scaling down a hot borate extraction method, a quick and easy method was developed to extract total RNA from ovaries of three genotypes ranging in ages from -1 to 10 days post anthesis (dpa) with an average yield of 600 µg/gfw (gram fresh weight) and ten dpa ovaries of 64 genotypes. The method was also successfully tested using cotton leaves and anthers. The quality and quantity of total RNA were suitable for reverse transcription-polymerase chain reaction (RT-PCR) and cDNA-amplified fragment length polymorphism (cDNA-AFLP) analysis.

              

Key words: Cotton, ovaries, RNA isolation, reverse transcription-polymerase chain reaction (RT-PCR), cDNA-amplified fragment length polymorphism (cDNA-AFLP).

Abbreviation

cDNA-AFLP, cDNA-amplified fragment length polymorphism; dpa,days post anthesis; RT-PCR, reverse transcription-polymerase chain reaction.