Comparison of Purity and Concentration Values of Mycobacterium tuberculosis DNA Extraction Result from the Boiling and Spin Column Method
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Abstract
The Polymerase Chain Reaction (PCR) technique is extensively employed in molecular biology to precisely detect Mycobacterium tuberculosis. Prior to conducting PCR, extracting of high-quality genomic Deoxyribonucleic Acid (DNA) is crucial to ensure accurate and reliable results. The primary objective of this study is to conduct a comparative analysis of the purity and concentration of M. tuberculosis DNA acquired through the utilization of the boiling method and the spin column extraction methods. A descriptive comparative research design was employed, utilizing a sample of 16 sputum specimens that had previously been confirmed as positive for M. tuberculosis through Acid-Fast Bacteria (AFB) examination and Molecular Rapid Test (MRT). The extraction of DNA was carried out using the boiling method and the spin column method. Subsequently, the concentration and purity of the extracted DNA were assessed using the NanoDrop Spectrophotometer, and the results were compared. The obtained yield of M. tuberculosis DNA isolates through the boiling method ranged from 9.6 ng/µL to 1258.7 ng/µL, with an average purity value of 1.23. Conversely, for the spin column method, the concentration of M. tuberculosis DNA isolates ranged from 8.7 ng/µL to 207.8 ng/µL, with an average purity value of 1.83. In conclusion, there is a significant difference between the purity and concentration of M. tuberculosis DNA extraction results using the boiling method and spin column methods.
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Copyright (c) 2023 Intan Febriyanti, Ai Djuminar, Fusvita Merdekawati, Asep Iin Nur Indra
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References
Maison DP. Tuberculosis pathophysiology and anti-VEGF intervention. J Clin Tuberc Other Mycobact Dis. 2022;27:100300. DOI: 10.1016/j.jctube.2022.100300 DOI: https://doi.org/10.1016/j.jctube.2022.100300
Iskandar D, Suwantika AA, Pradipta IS, Postma MJ, van Boven JFM. Clinical and economic burden of drug-susceptible tuberculosis in Indonesia: national trends 2017–19. Lancet Glob Heal. 2023;11(1):e117–25. DOI: 10.1016/S2214-109X(22)00455-7 DOI: https://doi.org/10.1016/S2214-109X(22)00455-7
World Health Organization. Global Tuberculosis report 2022 [Internet]. 2022. Available from: https://www.who.int/teams/global-tuberculosis-programme/tb-reports/global-tuberculosis-report-2022
Kementerian Kesehatan RI. Situasi TBC di Indonesia [Internet]. 2021 [cited 2022 Aug 5]. Available from: https://tbindonesia.or.id/pustaka-tbc/informasi/tentang-tbc/situasi-tbc-di-indonesia-2/
Jilani TN, Avula A, Gondal AZ, Siddiqui AH. Active Tuberculosis. In: NCBI bookshelf a service of the national library of medicine, national institutes of health [Internet]. StatPearls Publishing, Treasure Island (FL); 2023. p. 2. Available from: https://www.ncbi.nlm.nih.gov/books/NBK513246/?report=reader
MacPherson P, Lebina L, Motsomi K, Bosch Z, Milovanovic M, Ratsela A, et al. Prevalence and risk factors for latent tuberculosis infection among household contacts of index cases in two South African provinces: Analysis of baseline data from a cluster-randomised trial. PLoS One [Internet]. 2020;15(3):1–14. DOI: 10.1371/journal.pone.0230376 DOI: https://doi.org/10.1371/journal.pone.0230376
Babafemi EO, Cherian BP, Ouma B, Mogoko GM. Paediatric tuberculosis diagnosis using Mycobacterium tuberculosis real-time polymerase chain reaction assay: a systematic review and meta-analysis. Syst Rev [Internet]. 2021;10(1):1–14. DOI: 10.1186/s13643-021-01836-w DOI: https://doi.org/10.1186/s13643-021-01836-w
Fihiruddin, Ilmi HF, Khusuma A. Variations in boiling temperature in amplification of the M. tuberculosis inHA gene PCR method. [Variasi temperatur boiling pada amplifikasi Gen inHA M. tuberculosis Metode PCR]. Titian Ilmu J Ilm MultiS ciences. 2022;14(2):57–62. DOI: 10.30599/jti.v14i2.1661
García-Alegría AM, Anduro-Corona I, Pérez-Martínez CJ, Corella-Madueño MAG, Rascón-Durán ML, Astiazaran-Garcia H. Quantification of DNA through the nanodrop spectrophotometer: methodological validation using standard reference material and sprague dawley rat and human DNA. Int J Anal Chem. 2020;2020: 8896738. DOI: 10.1155/2020/8896738 DOI: https://doi.org/10.1155/2020/8896738
Bello GL, Morais FCL, Wolf JM, Gehlen M, Soares T dos S, Halon ML, et al. Improvement of Mycobacterium tuberculosis detection in sputum using DNA extracted by sonication. Brazilian J Infect Dis. 2020;24(5):398–404. DOI: 10.1016/j.bjid.2020.08.006 DOI: https://doi.org/10.1016/j.bjid.2020.08.006
Junior JCR, Tamanini R, Soares BF, Oliveira AM De, Silva FDG, Fernandes F, et al. Efficiency of boiling and four other methods for genomic DNA extraction of deteriorating spore-forming bacteria from milk. Semin Ciências Agrárias, Londrina. 2016;37(5):3069–78. DOI: 10.5433/1679-0359.2016v37n5p3069 DOI: https://doi.org/10.5433/1679-0359.2016v37n5p3069
Shi R, Lewis RS, Panthee DR. Filter paper-based spin column method for cost-efficient DNA or RNA purification. PLoS One. 2018;13(12):1–14. DOI: 10.1371/journal.pone.0203011 DOI: https://doi.org/10.1371/journal.pone.0203011
Hidayat MT, Prasetyo EN. Modified Spin Column-Based RNA Extraction Methods of Staphylococcus aureus using PureLink® RNA Mini Kit and basic laboratory instrument. Indones J Med Lab Sci Technol. 2021;3(2):73–80. DOI: 10.33086/ijmlst.v3i2.1863 DOI: https://doi.org/10.33086/ijmlst.v3i2.1863
Ndhlovu V, Mandala W, Sloan D, Kamdolozi M, Caws M, Davies G. Evaluation of the efficacy of two methods for direct extraction of DNA from Mycobacterium tuberculosis sputum. J Infect Dev Ctries. 2018;12(12):1067–72. DOI: 10.3855/jidc.10592 DOI: https://doi.org/10.3855/jidc.10592
European Centre for Disease Control. Handbook on tuberculosis laboratory diagnostic methods in the European Union [Internet]. Technical Report. 2018. 50–59 p. Available from: www.ecdc.europa.eu
Nasrollahzadehsabet M, Esmeilzadeh E, Shirmohammady N, Heidari MF. The Effect of EDTA buffer and temperature on DNA extraction from teeth for molecular forensic assessment. Ann Mil Heal Sci Res. 2021;19(2): e113043. DOI: 10.5812/amh.113043 DOI: https://doi.org/10.5812/amh.113043
Nurhayati B, Darmawati S. Cell and molecular biology. [Biologi Sel dan Molekuler]. Pusat Pendidikan SDM Kesehatan Badan Pengembangan dan Pemberdayaan Sumber Daya Manusia Kesehatan, Kemenkes RI.; 2017.
Afnan Uda MN, Parmin NA, Jambek AB, Hashim U, Uda MNA, Shaharuddin SNA. Evaluation and optimization of genomic DNA extraction from food sample for microfluidic purpose. IOP Conf Ser Mater Sci Eng. 2020;743(1). DOI: 10.1088/1757-899X/743/1/012031 DOI: https://doi.org/10.1088/1757-899X/743/1/012031
Chauhan T. 4 Important functions of nuclease-free water [Internet]. Genetic Education. 2022. Available from: https://geneticeducation.co.in/4-important-functions-of-nuclease-free-water/
Setiaputri AA, Rohmad Barokah G, Alsere Bardian Sahaba M, Dini Arbajayanti R, Fabella N, Mustika Pertiwi R, et al. Comparison of DNA isolation methods in fresh and processed fishery products. [Perbandingan metode isolasi DNA pada produk perikanan segar dan olahan]. J Pengolah Has Perikan Indones. 2020;23(3):447–58. DOI: 10.17844/jphpi.v23i3.32314 DOI: https://doi.org/10.17844/jphpi.v23i3.32314
Sophian A, Syukur A. Analysis of Purity and Concentration of Isolated DNA in making raw DNA of rat species. Erud Indones J Food Drug Saf. 2021;1(2):1–5. DOI: 10.54384/eruditio.v1i2.75 DOI: https://doi.org/10.54384/eruditio.v1i2.75
Dewanata PA, Mushlih M. Differences in DNA purity test using UV-Vis Spectrophotometer and Nanodrop Spectrophotometer in Type 2 Diabetes Mellitus patients. Indones J Innov Stud. 2021;15:1–10. DOI: 10.21070/ijins.v15i.553 DOI: https://doi.org/10.21070/ijins.v15i.553
Shen C-H. Quantification and analysis of nucleic acids. In: Diagnostic molecular biology (Second Edition) [Internet]. 2nd ed. Department of Biology, College of Staten Island, City University of New York, New York, NY, United States: Elsevier; 2023. Available from: https://www.sciencedirect.com/topics/biochemistry-genetics-and-molecular-biology/ultraviolet-spectrophotometry
Dayanti FG, Djuminar A, Dermawan A, Tantan A. Comparison of quantitative measurement values of Salmonella typhi DNA extraction results using the boiling method, NaOH, commercial kits. [Perbandingan Nilai pengukuran kuantitatif hasil ekstraksi DNA Salmonella typhi menggunakan metode boiling, NaOH, Kit komersial]. J Ris Kesehat Poltekkes Depkes Bandung. 2019;11(1):350–7. DOI: 10.34011/juriskesbdg.v11i1.762
Thermo Fisher Scientific Inc. NanoDrop 1000 Spectrophotometer V3 . 8 User ’ s Manual. 2012;11(1):10. Available from: www.nanodrop.com
Dorado G, Besnard G, Unver T, Hernández Molina P. Polymerase chain reaction (PCR). 2019;1–3(6):473–492. DOI: 10.1016/B978-0-12-801238-3.08997-2 DOI: https://doi.org/10.1016/B978-0-12-801238-3.08997-2
