Rapid and economical protocols for genomic and metagenomic DNA extraction from oak (Quercus brantii Lindl.)

Abstract

& Key message Two new efficient, fast and low-cost metagenomic DNA extraction methods were developed for different Persian oak tissues (leaf, stem, root, and rhizospheric) and soil samples. & Context The new "omics" studies on the genus Quercus are of importance to help finding efficient strategies for overcoming environmental challenges, and to do this, presence of efficient DNA extraction protocols for different Quercus species are very critical. & Aims The objective of the present study was to develop new efficient methods for extraction of metagenomic DNA (mDNA) from of Persian oak (Quercus brantii Lindl.) tissues. & Methods The efficiency of two newly developed mDNA extraction methods, including indirect SDS-based (ISB or concentrate method) and one spin column-based method (SCB) were compared to that of two classical direct methods, including CTABbased and SDS-based methods, and two commercial mDNA extraction kits. & Results The maximum average yield of mDNA for all samples (leaf, stem, root, bulk, and rhizospheric soils) was obtained by SCB (258 ng/μl) and ISB (189 ng/μl) methods, respectively. Successful PCR amplification for 16SrRNA and ITS sequence was consistently observed for ISB, SCB, and kit-extracted mDNAs, which confirmed the high purity of mDNA extracted by these methods. The new methods showed more than 96% quantitative PCR efficiency, and partial restriction digestion and metagenomic library construction confirmed the high efficiency of the newly developed methods. & Conclusion It could be concluded that two new protocols enhanced efficiency (yield, purity, and cost) of mDNA extraction from different tissues of Persian oak.

Olive Fungal Epiphytic Communities Are Affected by Their Maturation Stage

“…All steps for extracting DNA from olive epiphytes were performed at room temperature, except when clearly stated otherwise. The epiphytic genomic DNA was extracted through an adaptation of the Ahmadi et al protocol [17]. The pellet was suspended in 1 mL of phosphate buffer solution (1×, pH 6.7) and was continuously mixed for 3 min at 25 • C, at 160 rpm.…”

Versatile DNA extraction from diverse plant taxa using ionic liquids and magnetic ionic liquids: a methodological breakthrough for enhanced sample utility

“…Q. macrocarpa (an oak) is known to be a challenging plant genus for DNA extraction due to the presence of high levels of phenolic substances and secondary metabolites that are difficult to eliminate [36,37]. Inhibition of DNA amplification for Q. macrocarpa is likely due to either the co-extraction of polyphenolics and polysaccharides which can bind with DNA making it inaccessible to the polymerase enzyme or secondary metabolites that inhibit enzymatic activity [38][39][40]. Ethanol treatment can be a viable option in tissue preservation and removal of chlorophyll and secondary metabolites however, it may not be the ideal pretreatment method for plant taxa such as oaks.…”

Metagenomics: An Effective Approach for Exploring Microbial Diversity and Functions

“…DNA extraction should also be conducted carefully to limit any impurities from the host DNA. In previous studies, specific protocols were developed for different samples such as human fecal samples, tropical soils, and plant tissues [29][30][31]. Meanwhile, commercial kits for metagenomics have also been developed such as FastDNA™Spin Kit for Soil (MP Biomedicals, Irvine, CA, USA), Favor-Prep™ Soil DNA Isolation Mini Kit (Favorgen Biotech, Taiwan, China), MagAttract Pow-erSoil DNA KF Kit (Qiagen, Redwood, CA, USA), PureLink MicrobiomeTM Purification Kit (ThermoFisher Scientific, Waltham, MA, USA), and ZymoBIOMICS 96 Magbead DNA Kit (ZymoBIOMICS, Irvine, CA, USA).…”