M2 Metagenome: Hen manure and straw compost

Description

Functional metagenomes, by combining new generation sequencing technologies, bio-informatics and genetic engineering, make it possible, even in non-cultivable microbial consortia, to identify enzymes with new technological characteristics, to technologize them and use them in various processes in order to obtain products with "added value". DNA sample of the first phase compost microflora (based on straw and chicken manure, 75 oC) from Jraber, Armenia (40°20'16.7"N 44°39'19.3"E) was sequenced by long-read nanopore technology using a MinION Mk1B apparatus with R9․4.1 flow cell. A total of 327,921 reads were analyzed with an average of 3269 bases per reads. 98 % Bacteria, 1% Eukaryota and 1% Archaea+Viruses were detected among obtained reads. Representatives of the Deinococcus-Thermus filum were in the lead - 55.75 %, followed by Proteobacteria - 10.44% and Firmicutes - 6.64%. In the genus level Thermus was in majority – 55.66 %, followed by Escherichia – 4.26 % and Pseudomonas – 2.04 %. And in species level Thermus thermophilus composed 49.32 %, followed by Thermus parvatiensis – 4.73 % and Escherichia coli – 4.24 %. The resulting metagenome contains the necessary number of genes to obtain recombinant cellulases, hemicellulases, and lignin-modifying enzymes for lignocellulose degradation.

Steps to reproduce

Metagenomic sequencing was performed according to the recommendations of Oxford Nanopore Technologies (UK). ONT's proprietary MinKNOW software was used for raw data collection and basecalling. DNA library preparation, as well as priming and loading of the spot ON MinION R9.4.1 flow cell, was conducted following the ONT gDNA-sqk-lsk109 protocol (GDE_9063_v109_revAP_25May2022.pdf) available at https://nanoporetech.com/es/document/ gDNA-sqk-lsk109?format=versions. The EPI2ME platform (version MTE_1014_v1_revCB_11Apr2016), developed by Metrichor Ltd., a subsidiary of Oxford Nanopore Technologies, was used for taxonomic characterization of raw data (FASTQ dataset). The FastQ WIMP (What’s In My Pot) workflow was employed for this purpose. The software is available at: https://nanoporetech.com/document/epi2me?format=versions. Metagenome assembly was performed using Flye (version 2.9.1), a de novo genome assembler designed for long-read sequencing data. It is optimized for assembling high-error-rate reads into high-quality, polished contiguous sequences (contigs) [15,16]. The software is available at: https://github.com/ mikolmogorov/Flye /releases/tag/2.9.1. Metagenome annotation was performed using Prokka (version 1.14.5), a prokaryotic genome annotator that provides high-quality, standardized annotations [17]. The software is available at: https://github.com/tseemann/prokka/releases/tag/v1.14.5.

Categories

Funders

• State Committee of Science and Education of Republic of Armenia

  Grant ID: project № 21AG-2I090.

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