Abstract
Conventional RNA extraction methods have been shown to produce poor quality RNA with low yields from cotton tissues. We present a protocol for quick isolation of cotton RNA with high yield and good quality. By combining a borate extraction buffer developed by Wan and Wilkins (1994) with Qiagen RNeasy spin columns, plus proteinase K treatment during extraction, we produced RNA from 0 dpa (day postanthesis) cotton ovules at an average yield of 1000 μg/gfw (gram fresh weight). The poly A+ RNA derived from these RNA preparations was capable of efficient in vitro translation and reverse transcription. This method could also be applied to other cotton tissues, including leaves, 7 dpa fibres, 7 dpa ovules, petals, hypocotyls, and roots, to produce RNA at yields ranging from 300–1000 μg/gfw with little or no degradation.
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Abbreviations
- ESTs:
-
expressed sequence tags
- dpa:
-
days postanthesis
- gfw:
-
gram fresh weight
References
Chang S, Puryear J, and Cairney J (1993) A simple and efficient method for isolating RNA from pine trees. Plant Mol Biol Rep 11: 113–116.
Dolferus R, Jacobs M, Peacock WJ, and Dennis ES (1994) Differential interactions of promotor elements in stress responses of theArabidopsis Adh gene. Plant Physiol 105: 1075–1087.
Hasenfratz MP, Tsou CL, and Wilkins TA (1995) Expression of two related vacuolar H+-ATPase 16-kilodalton proteolipid genes is differentially regulated in a tissue-specific manner. Plant Physiol 108: 1395–1404.
Logemann J, Schell J, and Willmitzer L (1987) Improved method for the isolation of RNA from plant tissues. Anal Biochem 163: 16–20.
Maniatis T, Fritsch EF, and Sambrook J (1982) Gel electrophoresis of RNA. In: Molecular Cloning: A Laboratory Manual, pp 199–203, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York.
Murashige T and Skoog F (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant 15: 473–479.
Spencer D, Higgins TJV, Button SC, and Davey RA (1980) Pulse labelling studies on protein synthesis in developing pea seeds and evidence of a precursor form of legumin small subunit. Plant Physiol 66: 510–520.
Wan CH and Wilkins TA (1994) A modified hot borate method significantly enhances the yield of high-quality RNA from cotton (Gossypium hirsutum L.). Anal Biochem 223: 7–12.
Whittaker DJ and Triplett BA (1999) Gene specific changes in alpha-tubulin transcript accumulation in developing cotton fibres. Plant Physiol 121: 181–188.
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Wu, Y., Llewellyn, D.J. & Dennis, E.S. A quick and easy method for isolating good-quality RNA from cotton (Gossypium hirsutum L.) tissues. Plant Mol Biol Rep 20, 213–218 (2002). https://doi.org/10.1007/BF02782456
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DOI: https://doi.org/10.1007/BF02782456