| dc.description.abstract | DNA extraction is a critical step in molecular biology research, enabling the analysis of genetic material from various organisms. The development of efficient and reliable DNA extraction methods is essential for accurate downstream analyses and scientific advancements (Alberts et al., 2014). This project focused on investigating the yield and purity of DNA extracted from Eschericia coli (E. coli) and Mycobacterium tuberculosis (M. tuberculosis) bacteria, and comparing the performance of a home-made DNA extraction kit between these two bacterial species. The objectives of the study were to identify and evaluate the most effective combination of readily available reagents for developing a home-made DNA extraction kit for Eschericia coli and Mycobacterium tuberculosis bacteria samples, to determine the yield and purity of the DNA that was extracted from Eschericia coli and Mycobacterium tuberculosis bacteria, and to compare the performance of the home-made kit in the extraction of DNA between Eschericia coli and Mycobacterium tuberculosis bacteria. The DNA extraction procedure involved the use of a home-made kit comprising readily available reagents. E. coli and M. tuberculosis samples were subjected to cell lysis using a combination of Tris-HCL (pH 8.0), EDTA, SDS, and proteinase K. DNA purification was performed using sodium chloride and ethanol, followed by DNA washing with ethanol and elution in nuclease-free water. The extracted DNA was quantified using spectrophotometric analysis, and its purity was assessed by measuring the absorbance ratios (A260/A280). The amplifiability of the DNA was quantified using conventional PCR and Gel Electrophoresis. The results revealed significant differences in DNA yield and purity between E. coli and M. tuberculosis samples. E. coli consistently exhibited higher DNA yield and greater DNA purity compared to M. tuberculosis. The differences in DNA characteristics can be attributed to variations in cell wall composition and other biological factors specific to each bacterial species. The study also found out that the home-made kit was much cheaper compared to if a commercial kit was used, indicating the cost effectiveness of the home-made it. The study findings indicate that the performance of the home-made DNA extraction kit varied between E. coli and M. tuberculosis, with superior results obtained for E. coli samples. These findings highlight the importance of considering the specific characteristics of the target organisms when designing and optimizing DNA extraction protocols. The variations in DNA yield and purity between the two bacterial species have implications for future research involving these organisms. Based on the study findings, it is recommended to optimize the home-made DNA extraction kit for M. tuberculosis samples by adjusting the lysis buffer composition and proteinase K concentration. Exploring alternative reagents, such as lysozyme or lytic enzymes, specific to M. tuberculosis, can enhance cell lysis. Additionally, investigating alternative extraction methods, like bead-based or column-based systems tailored for mycobacterial DNA extraction, may improve results. Further understanding the impact of the unique M. tuberculosis cell wall composition on DNA extraction efficiency and exploring innovative approaches, such as specialized lysis buffers or targeted enzymes, can overcome challenges associated with extracting DNA from gram-positive bacteria. Implementing these recommendations will enhance the kit's performance, improving DNA yield, purity, and amplifiability for better molecular research on M. tuberculosis. It is also recommended to further investigate the impact of the bacterial cell wall structure on DNA extraction efficiency and explore innovative approaches for overcoming the challenges associated with extracting DNA from gram-positive bacteria like M. tuberculosis. | en_US |
