Fast protocol for extraction of DNA from Prosopis spp leaves (plant adapted to arid environment) without liquid nitrogen

Abstract

ABSTRACT. The extraction of high-quality genomic DNA from Prosopis spp for polymerase chain reaction (PCR) amplification is complicated, owing to the presence of a high percentage of secondary metabolites that bind to or co-precipitate with nucleic acids. In the present study, we report a modified sodium dodecyl sulfate/phenol protocol that eliminates the use of liquid nitrogen in the maceration process, β-mercaptoethanol in the buffer extraction, and the ethanol precipitation step. The A 260 /A 280 absorbance ratios of the isolated DNA were approximately 2.0 to 1.9, suggesting that the DNA fraction was pure and can be used for further PCR analysis. The DNA isolated by this protocol is of sufficient quality for molecular applications; this technique could be applied to other organisms that have similar substances that hinder DNA extraction. Finally, this proposal represents an alternative fast, cheap, and effective method for the isolation of genomic DNA from fresh leaves of Prosopis spp, even in low-technology laboratories.

Characterization and selection of markers associated with resistance analogous genes as input for genetic analysis of Prosopis juliflora (Sw.) DC.

“…However, such use raises the procedure costs. Therefore, in the present study, the absence liquid nitrogen did not interfere with the quality of the extracted DNA, offering good results, which are also in agreement with the literature that advocates a low cost technique with fewer steps in the procedure (Michel-López et al, 2013;Santos & Araújo, 2017).…”

Análise comparativa de protocolos para extração de DNA genômico em Prosopis juliflora (Sw.) DC

“…Outros estudos corroboram esses resultados, demostrando não haver necessidade da utilização do β-mercaptoetanol na extração de DNA (SCHMITT et al, 2014;SILVA et al, 2014 de PCR. Outros estudos reportaram que a ausência de nitrogênio líquido no processo de maceração torna custo operacional menos oneroso (MICHEL-LÓPEZ et al, 2013;DOS SANTOS;ARAÚJO, 2017).…”

“…Embora a espécie possua muito potencial de uso pelo ser humano e estudos de diveridade genética sejam o ponto de partida para se efetivar ações de conservação e de melhoramento de espécies com potencial econômico (LOPES et al, 2011;BESSEGA et al, 2018;FALEIRO et al, 2018), pouco se conhece quanto à diversidade genética de P. juliflora. Para estudos dessa natureza, faz-se necessária a obtenção de ácidos nucleicos, em quantidades e qualidades que possibilitem a amplificação do material genômico e a seleção de iniciadores potenciais (MICHEL-LÓPEZ et al, 2013). O processo de extração de DNA é parte fundamental em estudos moleculares, sendo sua eficiência diretamente relacionado à pureza, à quantidade e à qualidade do material genômico obtido (DA ROCHA et al, 2017).…”

Comparative Analysis of the Complete Chloroplast Genome of Three Prosopis Species in Jordan

“…The DNA extraction of chloroplasts, nucleic acid quality control, quantification, and gel electrophoresis were conducted to prepare the library for quantification using a bio-analyzer (37). DNA was extracted from plant tissues by using a modified protocol (45) used at the Biotechnology Omics Lab and the University of Nizwa, Oman.…”