Ligninolytic enzymes improve soil DNA purity: Solution to methodological challenges of soil metagenomics

• Published in: Journal of molecular catalysis. B, Enzymatic Vol. 83; pp. 73 - 79
• Main Authors: Sharma, Krishna Kant, Sharma, Sonia, Karp, Matti, Kuhad, Ramesh Chander
• Format: Journal Article
• Language: English
• Published: Amsterdam Elsevier B.V 01.11.2012; Elsevier
• Subjects: Bioconversions. Hemisynthesis; Biological and medical sciences; Biotechnology; Fundamental and applied biological sciences. Psychology; Humic acid; Ligninolytic enzymes; Metagenomic DNA; Methods. Procedures. Technologies; Phenolic compounds; Soil; Soil; Phenolic compounds; Metagenomic DNA; Ligninolytic enzymes; Humic acid; Soils; Metagenomics; Enzyme; DNA; Phenols; Ligninolytic enzyme; Optimization
• ISSN: 1381-1177; 1873-3158
• DOI: 10.1016/j.molcatb.2012.07.010

[Display omitted] ► First report of ligninolytic enzymes as purifying agent for soil metagenomic DNA. ► DNA amenable for molecular manipulations is isolated using this protocol. ► Purity of isolated DNA was checked using restriction digestion, PCR and qPCR. ► Yield and purity of the DNA isolated was higher as compared to existing protocols. A new, improved and cost effective protocol for extraction of pure, high molecular weight metagenomic DNA from soil samples using ligninolytic enzymes has been developed. DNA yield of 17μgg−1 of soil from termite nest soil and 25μgg−1 from forest soil and size of 15–20kb was achieved by the optimized protocol. More than 90% of DNA was recovered after purification and there was 81% reduction in humic acid content. The strategy allowed processing of several samples at a given time and yielded purified DNA. The isolated metagenomic DNA sample could be readily processed for restriction digestion and PCR amplification. The quantitative PCR revealed CP value of 21 for the treated samples whereas the untreated samples could not cross the threshold fluorescence even after 45 cycles. This clearly indicated that the isolated metagenomic DNA is amenable for molecular manipulations.