Optimization of microbial DNA extraction from Dendrocalamus asper (buluh betung) for metagenomic sequencing.

• Main Author: Yamunaa d/o Sangaya
• Format: Undergraduate Final Project Report
• Language: English
• Published: 2017
• Online Access: http://discol.umk.edu.my/id/eprint/7256/1/YAMUNAA%20DO%20SANGAYA.pdf; http://discol.umk.edu.my/id/eprint/7256/

Isolation of genomic DNA of microbial cells in the woody plants presents a special challenge to molecular work especially when several plant metabolites such as phenolic compound that have chemical properties similar to those of nucleic acids which cause these compounds to co-precipitate with DNA and RNA thus resulting in low quantity and quality of the nucleic acid. In view of the fact that none of the published protocol has established itself as universally applicable to extract DNA of plant microbes from all plant varieties, the commonly used techniques require adaptation before it can be used with plant samples. In this work, attempts were made to extract a good quality DNA from woody sample of Dendrocalamus asper (Buluh Betung) using different protocols including conventional and extraction kit involving the use of Cetyltrimethylammonium Bromide ( CTAB) and Sodium Dodecyl Sulphate (SDS) buffers together with Wizard Genomic DNA purification Kit. Apart from that, the types of chemicals beside the external factors that have significant effects on the condition of the extracted were also emphasized in this review. The extracted genomic DNA was eventually subjected to agarose gel electrophoresis and spectrophotometer to determine its yield and purity levels. In comparison of all the four different protocol, it was found that the carefully modified CTAB conventional method generally produces a consistent and higher yield of DNA with a concentration of 19.66 ng/μL in relative to only 8.90 ng/μL of DNA concentration for DNA extracted using SDS-based DNA extraction buffer as well as the extraction kit. Therefore, the CTAB- based extraction buffer method was proven to be reliable in generating a good quality of DNA from these particular genera.