Construction of Cosmid Library of Metagenome Derived from Compost Sample and the Improvement of DNA Isolation Techniques

Abstract:

Abundant humic acid present in compost hinder the metagenomic study of uncultured microorganisms in compost environment. We developed an effective method for isolating metagenomic DNA from compost samples. By combinations of Sephadex G200 + acid washed PVPP column and electroelution techniques, this method was effective in purifying of metagenomic DNA from compost samples. With this purified DNA, we succeeded in constructing a cosmid library containing about one hundred thousands clones. And a novel β-glucosidase gene was cloned and sequenced from this library. Targeting the problem of low frequency of positive clones in the library, we also studied the effects of different centrifugal speeds on the content of contaminated eukaryotic DNA in the isolated metagenomic DNA, in order to preclude the presence of eukaryotic DNA in metagenomic library.