Comparison of Three Different DNA Isolation Methods To Degradate The Trichoderma Fungi Cell Wall

Abstract

Trichoderma fungi cell walls are composed of chitin compounds that are very sturdy and resistant to enzyme activity. The specific techniques and methods are needed to degradate of fungi cell wall. This study aims to analyze the best DNA isolation method in lysing fungal cell walls by comparing several isolation methods. The methods are using DNeasy Plant Mini Kit, DNeasy Plant Mini Kit combination with heating, and DNeasy Plant Mini Kit combination in a physical way (grinding the sample using mortar and pestle in liquid nitrogen). This research was conducted in March to September 2018 at the Microbiology Laboratory, Laboratory of Biotechnology and Genetics, as well as the Integrated Research Laboratory, FMIPA, UNP. Data were analyzed qualitatively by agarose gel electrophoresis and quantitatively by calculating the purity and concentration of DNA using Nanodrop. The results showed that the DNeasy Plant Mini Kit combination method by physical means gave the highest concentration of DNA isolation of 2.4 μg / ml with a purity value of 1.857. In the DNeasy Plant Mini Kit method and the DNeasy Plant Mini Kit combination method with heating, the DNA concentrations obtained were 1.211 µg / ml and 0.933 µg / ml respectively with a purity value of 1.728 and 1.708. The results of the electrophoresis test showed a thin band of DNA bands in samples isolated by a combination of kit methods and physical methods, whereas in the other two samples no DNA bands were found. It can be concluded that the method of DNA isolation that can be used as a reference to degrade Trichoderma mushroom cell walls is a combination of kit and liquid nitrogen method with a note that additional levels of isolates are needed.