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Abstract
This study aimed to detect Escherichia coli directly without DNA extraction. The nucleus membrane and cell membranes of the Escherichia coli are composed of a phospholipid bilayer, damaged if heated at 950C. Pre-denaturation and denaturation of PCR were carried out at 950C. The two stages are thought to break down the Escherichia coli cells, so that the DNA that comes out of the cells can directly become a template in the PCR analysis. In this study, PCR analysis was carried out using Escherichia coli culture, Escherichia coli bacteria culture incubated at 950C, and Escherichia coli bacteria cultures incubated at 650C + on ice as templates. The results showed that PCR analysis using Escherichia coli culture directly and Escherichia coli culture incubated at 650C + on ice as templates produced very thin DNA bands with a size of 580 bp. while PCR analysis using Escherichia coli bacteria culture incubated at 950C as a template produced thick DNA bands with a size of 580 bp. This study's results are very useful for saving time and costs in the detection of Escherichia coli bacteria. The sample to be tested does not need DNA isolation as usual, but only needs to be incubated at 950C for 10 minutes.
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Folia Medica Indonesiana is a scientific peer-reviewed article which freely available to be accessed, downloaded, and used for research purposes. Folia Medica Indonesiana (p-ISSN: 2541-1012; e-ISSN: 2528-2018) is licensed under a Creative Commons Attribution 4.0 International License. Manuscripts submitted to Folia Medica Indonesiana are published under the terms of the Creative Commons License. The terms of the license are:
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References
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References
Bernardi B, Kayacan Y, Akan M, et al (2019). Overexpression of rad51 enables pcr-based gene targeting in lager yeast. Microorganisms 7, 1-12.
Cherpillod P, Schibler M, Vieille G, et al (2016). Ebola virus disease diagnosis by real-time RT-PCR: A comparative study of 11 different procedures. Journal of Clinical Virology 77, 9-14.
El-Ashram S, Al Nasr I, Suo X (2016). Nucleic acid protocols: Extraction and optimization. Biotechnology Reports 12, 33-39.
Farrar JS, Wittwer CT (2015). Extreme PCR: Efficient and specific DNA amplification in 15-60 seconds. Clinical Chemistry 61, 145-153.
Ghatak S, Muthukumaran RB, Nachimuthu SK (2013). A simple method of genomic DNA extraction fromhuman samples for PCR-RFLP analysis. J Biomol Tech 24, 224-231.
Jackson M, Gradmann C (2018). Medical bacteriology: The routledge history of disease. Routledge, UK.Lorenz TC (2012). Polymerase chain reaction: Basic protocol plus troubleshooting and optimization strategies. Journal of Visualized Experiments 63, 1-15.
Reece JB, Lisa AU, Peter VM, et al (2010). Campbell biology. Pearson, London.
Sambrook J, Russell DW (2001). Molecular Cloning: A Laboratory Manual, Third Edition. Cold Spring Harbor Laboratory Press, New York.
Setiati N, Partaya, Hidayah N (2019). The use of two pairs primer for CO1gene amplification on traded stingray at fish auction Tasik Agung Rembang. Journal of Physics: Conference Series 1567, 1-6.
Tsen HY, Lin CK, Chi WR (1998). Development and use of 16S rRNA gene targeted PCR primers for the identification of Escherichia coli cells in water. Journal of Applied Microbiology 85, 554-60.
Verma SC, Qian Z, Adhya SL (2019). Architecture of the Escherichia coli nucleoid. PLoS Genet 15, 1-35.
Widodo WT, Yudianto A, Astuti WSP (2018). Identification of human DNA in mixture of human and chicken blood using PCR with specific primer of cytochrome B gene. Folia Medica Indonesiana54, 184-188.
Wu JY, Jiang XT, Jiang YX, et al (2010). Effects of polymerase, template dilution and cycle number on PCR based 16 S rRNA diversity analysis using the deep sequencing method. BMC Microbiology 10, 1-7.
Yustinadewi PD, Yustiantara PS, Narayan I (2018). Teknik perancangan primer untuk sekuen Gen MDR-1 varian 1199 pada sampel buffy coatpasien anak dengan LLA. JURNAL METAMORFOSA V, 105-111.