The isolation of high-quality plant genomic DNA is a major prerequisite in many plant biomolecular analyses. The presence of rigid cell walls and high levels of secondary metabolites often makes lysis and extraction of DNA from plant tissue difficult and challenging. Conventional plant cell lysis and DNA extraction methods require time-consuming sample preparation steps that often incorporate or generate numerous quantitative polymerase chain reaction (qPCR) inhibitors, which can limit their applicability in high throughput applications. In order to address these challenges, consolidated approaches combining rapid lysis and DNA extraction steps have the potential to shorten analyses while also eliminating unwanted contamination. We seek to exploit the unique properties of ionic liquids (ILs) and deep eutectic solvents (DESs) as solvents in the extraction of high-quality plant genomic DNA to provide high yields suitable for downstream applications, such as qPCR. In this study, a simple, fast, sustainable and environmentally-friendly extraction method is developed for the extraction of genomic DNA from Arabidopsis Thaliana plant tissue. The IL is directly applied to the plant tissue and the DNA enriched IL is recovered for qPCR analysis as opposed to conventional DNA extraction methods that involve harsh conditions, such as toxic chemicals that could be more destructive to the plant tissue. Optimization of the DNA extraction method in terms of solvent volume, incubation temperature, extraction time, and sample pretreatment methods are presented. The trihexyl(tetradecyl)phosphonium bis(trifluoromethylsulfonyl)imide ([P6,6,6,14+][NTf2-]) IL exhibits promise due to its high DNA extraction efficiency and compatibility with qPCR. Furthermore, the applicability of other ILs and DESs will be discussed. The DNA extraction methods using these solvents will also be applied to understand the relationship between the developmental stage of the plant and the recovery of DNA.

Rapid plant cell lysis and DNA extraction from Arabidopsis Thaliana using ionic liquids

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