Abstract

DNA Extraction from silica gel preserved common bean (Phaseolus vulgaris L.) leaves

Extraction of high-quality DNA from field specimen requires collection under liquid nitrogen which is not readily available in resource constrained laboratories in low- and middle-income countries (LMICs). A method of extracting DNA from silica gel preserved common bean (Phaseolus vulgaris L.) leaves is presented. Our method which does not involve the use of phenol, chloroform or isoamyl alcohol also obviates the need for low temperature incubation during the DNA extraction steps and the grinding of desiccated leaf tissue in liquid nitrogen. It relies on inactivating proteins using SDS and proteinase K and precipitation of polysaccharides using a high salt solution. DNA is further purified by exploiting its insolubility in aqueous media. We were able to extract high quality pure DNA (mean concentration 2.98 ± 0.84 μg/g of leaf tissue) with mean A260/280 of 2.1 ± 0.1 and A260/230 of 2.4 ± 0.15. The DNA was also found to be amenable to amplification using molecular marker types routinely used in molecular biology laboratories like random amplified polymorphic (RAPD) markers, inter simple sequence repeat (ISSR) markers, sequence-characterized amplified region (SCAR) markers and simple sequence repeat (SSR) markers. Our findings show that it is possible to obtain high quality DNA from leaf tissue preserved in silica gel. Our method will be invaluable to resource constrained laboratories especially in LMICs that cannot afford to buy or access liquid nitrogen in order to extract high quality DNA and to research groups undertaking field surveys that require several days or weeks off station without laboratory freezers to maintain the integrity of the tissues which is crucial for obtaining high quality DNA.

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