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. 2017 Jul;7(3):182.
doi: 10.1007/s13205-017-0846-y. Epub 2017 Jun 29.

Rapid and efficient method to extract metagenomic DNA from estuarine sediments

Kashif Shamim  1 , Jaya Sharma  1 , Santosh Kumar Dubey  2
Affiliations

Rapid and efficient method to extract metagenomic DNA from estuarine sediments

Kashif Shamim et al. 3 Biotech. 2017 Jul.

Abstract

Metagenomic DNA from sediments of selective estuaries of Goa, India was extracted using a simple, fast, efficient and environment friendly method. The recovery of pure metagenomic DNA from our method was significantly high as compared to other well-known methods since the concentration of recovered metagenomic DNA ranged from 1185.1 to 4579.7 µg/g of sediment. The purity of metagenomic DNA was also considerably high as the ratio of absorbance at 260 and 280 nm ranged from 1.88 to 1.94. Therefore, the recovered metagenomic DNA was directly used to perform various molecular biology experiments viz. restriction digestion, PCR amplification, cloning and metagenomic library construction. This clearly proved that our protocol for metagenomic DNA extraction using silica gel efficiently removed the contaminants and prevented shearing of the metagenomic DNA. Thus, this modified method can be used to recover pure metagenomic DNA from various estuarine sediments in a rapid, efficient and eco-friendly manner.

Keywords: Gene cloning; Metagenomic library; Metagenomics; PCR amplification; Restriction digestion; Spectrophotometry.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Fig. 1
Fig. 1
(A) Agarose gel analysis of metagenomic DNA isolated from estuarine samples. Lane 1–2 DNA isolated using present method from sample GUMD1. Lane 3–4 DNA isolated using present method from sample GUMD2. Lane 5–6 DNA isolated following the method of Amorim et al. (2008) from sample GUMD1 and GUMD2. Lane 7–8 DNA isolated following the method of Bürgmann et al. (2001) from sample GUMD1 and GUMD2. Lane 9–10 DNA isolated following the method of Yeates et al. (1997) from sample GUMD1 and GUMD2. Lane 11–12 DNA isolated following the method of Zhou et al. (1996) from sample GUMD1 and GUMD2. Lane 13 DNA isolated from cultural bacteria as a control. (B) Lane 1 DNA isolated following the method of Amorim et al. (2008) from sample GUMD3. Lane 2 DNA isolated following the method of Bürgmann et al. (2001) from sample GUMD3. Lane 3 DNA isolated following the method of Yeates et al. (1997) from sample GUMD3. Lane 4 DNA isolated following the method of Zhou et al. (1996) from sample GUMD3. Lane 5 DNA isolated following the present method from sample GUMD 3
Fig. 2
Fig. 2
Agarose gel analysis of Sau3AI digested metagenomic DNA. Lane 1–3 Sau3AI digested metagenomic DNA from estuarine sediments of GUMD1, GUMD2 and GUMD3, respectively
Fig. 3
Fig. 3
PCR amplification of 16S rDNA using recovered metagenomic DNA samples. Lane 1 1 Kbps DNA marker; Lane 2–4 16S rDNA amplicon
See this image and copyright information in PMC

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