Mycelium Lab Report

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9 MOLECULAR TAXONOMY AND EXPERINMENTAL SPECIES 9.1 INTRODUCTION The Molecular phylogenetics uses the structure and function of molecules and how they change over time to infer these evolutionary relationships. This branch of study emerged in 20th century but didn’t begin in earnest until the 1960s, with the advent of protein sequencing, PCR, electrophoresis, and other molecular biology techniques. Over the past 30 years, as computers have become more powerful and more generally accessible, researchers have been able to tackle the immensely complicated stochastic and probabilistic problems that define evolution at the molecular level more effectively. (Durbin et al., 1998; Hall, 2004; Li, 1997; Lio and Goldman, 1998; Patthy and Laszlo, 1998).Phylogenetic…show more content…
The fungal culture was grown in 100 ml PDB for 5 days. The culture was harvested and the mycelial mat was separated by filtration using Whatman No. 1 filter paper. Then mycelium was ground separately in pestle and mortar using liquid nitrogen. About 50 mg of the powdered mycelium was transferred into a microtube contained 500 µl of TES (100 mMTris, pH 8.0, 10 mM EDTA, 2% SDS). To which, 50 µg proteinase K was added and incubated for 1 h at 60°C with occasional gentle mixing. To the above mixture, 140 µl of 5 M NaCl was added to adjust the salt concentration to 1.4 M. Then 65 µl of 10% CTAB (CetylTrimethyl ammonium bromide) was added and incubated for 10 min at 65°C. To the above mixture, 700 µl of Chloroform and isoamyl alcohol (24:1) was added, mixed gently, incubated for 30 min at 0°C and centrifuged at 10000 rpm for 10 min at 4°C. The supernatant was transferred to a 1.5 ml tube; to which 225 µl of 5 M NH4Ac was added, mixed gently, incubated on ice for 30 min and centrifuged at 10,000 rpm for 15 min at 4°C. Then the supernatant was transferred to a fresh tube, 510 µl of isopropanol was added to precipitate the DNA and centrifuged immediately for 10 min at 10,000 rpm. The supernatant was removed; the pellet was washed twice with cold 70% ethanol, air-dried and suspended in 50 µl TE

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