Comparative assessment of methods for metagenomic DNA isolation from soils of different crop growing fields

Abstract

The isolation of good quality metagenomic DNA from diverse soil, in appreciable amount, is a prerequisite for metagenomics. The availability of commercial kits for isolation of genomic DNAs from soil has drastically expedited the application of metagenomics approach for identifying novel sources of industrially important enzymes. The quantitative and qualitative assessment of metagenomic DNA isolated using either the manual method or the kit-based method should be performed prior to its use in downstream applications. The metagenomic DNA isolated from six different soil samples, using three methods, were analyzed in terms of yield, quality and downstream application as template for PCR amplification. The yield of DNA was approximately 3.52, 7.35, and 232.42 μg of DNA per gram of soil sample for the kit-based method, kit-modified method, and manual method, respectively. The manual method seems to be promising based on better yield and lesser humic acid content than the other two methods. The maximum yield was obtained in the soil collected from teak forest with all the three methods, indicating maximum microbial content and diversity. Furthermore, in terms of its suitability as template DNA for PCR amplification using 16S RNA primer, all methods are equally well. Thus, comparative assessment of three methods revealed suitability of manual method based on DNA yield and humic acid content, which could be important for many downstream applications like library preparations during metageomics approach.

Peer Review #1 of "Towards optimized viral metagenomes for double-stranded and single-stranded DNA viruses from challenging soils (v0.1)"

“…2B; for proteins and phenol contamination; Maniatis et al 1982) and A260:230 ratios ( Fig. S2; for carbohydrates and phenols; Maniatis et al 1982;Tanveer, Yadav, and Yadav 2016). We posited that A260:280 is a more robust predictor of virome success, since previous work showed that A260:230 of DNA extracts had limited correlation to amplification success (Costa et al 2010;Ramos-Gómez et al 2014), although both are highly variable for low DNA concentrations typical for soil viral extracts.…”

Metagenomic applications in exploration and development of novel enzymes from nature: a review

“…Some researchers use different methods, depending on the type of research sample used [10]. Tanveer et al [27] have carried out DNA extraction of the metagenome using commercial kits and standard protocols. Metagenomic DNA extraction using commercial kits is the easiest method because it only uses chemicals that have been provided by the manufacturer.…”

“…However, some studies that use standard protocols show better results when compared to kits. Tanveer et al [27] tried to compare the extraction of metagenomic DNA from the soil using the HiPurA soil DNA isolation kit (Himedia) and standard protocol. The results revealed that the standard protocol produced the highest concentration of DNA.…”

CTAB influenced differential elution of metagenomic DNA from saltpan and marine sediments

“…As the physico-chemical properties of soil and sediment samples vary worldwide, it is difficult to develop a single, ideal total community DNA extraction protocol (Felczykowska et al 2015). A number of chemical, mechanical, and enzymatic methods and commercial kits have been developed to encounter this problem (Tsai and Olson 1991;Canto-Canche et al 2013;Tian et al 2013;Gutierrez-Lucas et al 2014;Devi et al 2015;Tanveer et al 2016). Moreover, several cell lysis methods and purification steps have been recommended for the efficient recovery of high quality metagenomic DNA.…”