Quality Improvement of the DNA extracted by boiling method in Gram negative bacteria

Omar B. Ahmed*, Anas S. Dablool

Abstract


Several methods of Deoxyribonucleic acid (DNA) extraction have been applied to extract bacterial DNA. The amount and the quality of the DNA obtained for each one of those methods are variable. The study aimed to evaluate bacterial DNA extraction using conventional boiling method followed by alcohol precipitation. DNA extraction from Gram negative bacilli was extracted and precipitated using boiling method with further precipitation by ethanol. The extraction procedure performed using the boiling method resulted in high DNA yields for both E. coli and K. pneumoniae bacteria in (199.7 and 285.7μg/ml, respectively) which was close to control method (229.3 and 440.3μg/ml). It was concluded that after alcohol precipitation boiling procedure was easy, cost-effective, and applicable for high-yield quality of DNA in Gram-negative bacteria.

Keywords


DNA; boiling method; extraction; Gram negative

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References


da Silva G A, Bernardi T L, Schaker P D C, Menegotto M, Valente P. Rapid Yeast DNA Extraction by Boiling and Freeze-Thawing Without Using Chemical Reagents and DNA Purification. Braz. arch. biol. Technol, 2012, 55(2), 319-327.

Ahmed OB, Asghar AH, Elhassan MM. Comparison of three DNA extraction methods for polymerase chain reaction (PCR) analysis of bacterial genomic DNA. Afr J Microbiol Res, 2014, 8(6), 598-602.

Reischl U, Linde HJ, Metz M, Leppmeier B, Lehn N. Rapid identification of methicillin-resistant Staphylococcus aureus and simultaneous species confirmation using real-time fluorescence PCR. J. Clin. Microbiol, 2000, 38, 2429-2433.

Queipo-Ortuño Ml, Colmenero JDD, Macias M, Bravo MJ, Morata P. Preparation of Bacterial DNA Template by Boiling and Effect of Immunoglobulin G as an Inhibitor in Real-Time PCR for Serum Samples from Patients with Brucellosis. Clin. Vaccine Immunol. 2008, 15(2), 293-296.

Araújo WL, de Angelis DA, Azevedo JL Direct RAPD evaluation of bacteria without conventional DNA extraction. Braz Arch Biol Technol, 2004, 47, 375-380.

Deak T, Chen J, Beuchat LR, Molecular characterization of Yarrowia lipolytica and Candida zeylanoides isolated from poultry. Appl Environ Microbiol, 2000, 66 (10): 4340-4344.

Fregel R, González A, Cabrera V M. Improved ethanol precipitation of DNA. Electrophoresis. 2010, 31, 1350-1352.

Moore E, Arnscheidt A, Kruger A, Strompl C, Mau M. Simplified protocols for the preparation of genomic DNA from bacterial cultures. Molecular Microbial Ecology Manual, Second Edition. 2004, 101, 3-18.

Ahmed OB, Omar AO, Asghar AH, Elhassan MM. Prevalence of TEM, SHV and CTXM genes in Escherichia coli and Klebsiella spp Urinary Isolates from Sudan with confirmed ESBL phenotype. Life Sci J, 2013, 10(2), 191-195.

An R, Jia Y, Wan B , Zhang Y, Dong P , Li J, Liang X. Non-Enzymatic Depurination of Nucleic Acids: Factors and Mechanisms. PLoS One, 2014, 9(12), e115950.

Rantakokko-Jalava K and Jalava J Optimal DNA Isolation Method for Detection of Bacteria in Clinical Specimens by Broad-Range PCR J Clin Microbiol, 2002, 40(11), 4211–4217.

Peng X, Yu K-Q, Deng G-H, Jiang Y-X, Wang Y, Zhang G-X, Zhou H-W. Comparison of direct boiling method with commercial kits for extracting fecal microbiome DNA by Illumina sequencing of 16S rRNA tags. Journal of Microbiological Methods, 2013, 95(3), 455-462.

Yamagishi J, Sato Y, Shinozaki N, et al. Comparison of Boiling and Robotics Automation Method in DNA Extraction for Metagenomic Sequencing of Human Oral Microbes. Alekseyenko AV, ed. PLoS ONE. 2016, 11(4), e0154389




DOI: http://dx.doi.org/10.21746/ijbio.2017.04.004

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