Korean Journal of Microbiology (2020) Vol. 56, No. 1, pp. 74-76 pISSN 0440-2413
DOI https://doi.org/10.7845/kjm.2020.9154 eISSN 2383-9902
Copyright ⓒ 2020, The Microbiological Society of Korea
Complete genome sequence of Bacillus velezensis SRCM102755, a high menaquinone-7 producer, isolated from Doenjang
Gwangsu Ha , Sua Im , Su-Jin Shin , Minji Ryu , JinWon Kim , Seong-Yeop Jeong , and Do-Youn Jeong*
Microbial Institute for Fermentation Industry (MIFI), Sunchang 56048, Republic of Korea
된장에서 분리한 menaquinone-7 고생성균주인
Bacillus velezensis SRCM102755 균주의 유전체 해독
하광수 ･ 임수아 ･ 신수진 ･ 류민지 ･ 김진원 ･ 정성엽 ･ 정도연*
(재)발효미생물산업진흥원
(Received Received December 18, 2019; Revised February 10, 2020; Accepted February 13, 2020)
*For correspondence. E-mail: [email protected];
Tel.: +82-63-653-9579; Fax: +82-63-653-9590
Bacillus velezensis SRCM102755, isolated from Doenjang was screened as a starter for Cheonggukjang containing high menaquinone-7 (MK-7) production. Here, we report the complete genome sequence of B. velezensis SRCM102755. The complete genome was sequenced by Pacific Biosciences (PacBio) single- molecule real-time (SMRT) RS II platform. The genome consists of a single, circular chromosome of 4,044,843 base pairs with a G + C content of 46.34%, and included 3,856 protein coding genes and 113 RNA genes (27 rRNA genes, 86 tRNA genes).
The genome was identified to contain at least of nine genes involved in MK-7 biosynthesis.
Keywords: Bacillus velezensis, complete genome sequence, menaquinone-7
Menaquinone-7 (MK-7) a highly bioactive homologue of vitamin K group has an important role in preventing diseases such as cardiovascular and bone diseases besides its positive impacts on blood clots (Theuwissen et al., 2012). As these diseases increase, researchers seek alternative approaches to preventing these health issues (Raspini and Cazzola, 2015).
Several studies showed that risk of bone fractures and cardio- vascular disorders by reducing the calcification in arteries is
significantly reduced by MK-7 consumption (Truong and Booth, 2011; Puri et al., 2015). The effective therapeutic dose of MK-7 for treatment of osteoporosis or cardiovascular diseases from MK-7 abundant dietary supplements such as fermented soybean, blue cheese and meat require the consume of impractical quantities. Therefore, development of a rich source of MK-7 as dietary supplements is highly required (Berenjian et al., 2012).
The Gram-positive, rod-shaped, and endospore-forming strains of the Bacillus group are the most characterized organisms for application to various industries (Parvathi et al., 2009).
Especially, Bacillus strains are used for production of many valuable components including MK-7 (Berenjian et al., 2012;
Xu et al., 2017). In this report, we present complete genome sequence of B. velezensis SRCM102755 as a MK-7 producer in Cheonggukjang (unpublished data). Strain SRCM102755 was isolated from Doenjang produced in Sunchang, Republic of Korea (35.374384 N 127.137571 E) using serial dilution method.
When 16S ribosomal RNA gene was compared to other members of the Bacillus group using EzBioCloud database, the most similar strains to strain SRCM102755 were Bacillus velezensis AY603658 (99.93%), Bacillus subtilis subsp. subtilis (99.80%).
The genomic DNA of strain SRCM102755 was prepared
according to the protocol in DNeasy Blood & Tissue Kits
Complete genome sequence of B. velezensis SRCM102755 ∙ 75
Korean Journal of Microbiology, Vol. 56, No. 1 (QIAGEN). DNA concentration was estimated by picogreen
(Thermo Fisher Scientific) using Victor 3 fluorometer (PerkinElmer) and purity was determined by Nanodrop ONE (Termo scientific).
The complete genome of strain SRCM102755 was sequenced using a Pacific biosciences (PacBio) RS II single-molecule real-time (SMRT) sequencing system with a 20 kb SMRTbell
TMtemplate at Macrogen Inc. The de novo assembly was performed by Hierarchical Genome Assembly (HGAP) version 3 within SMRT Portal version 2.3 (Chin et al., 2013). Correcting bases, fixing mis-assemblies and filling gaps were carried out using Pilon version 1.21 (Walker et al., 2014). Prediction of gene and functional annotation were identified using the NCBI Prokatyotic Genomes Automatic Annotation Pipeline (PGAAP) (http://www.
ncbi.nlm.nih.gov/genome/annotation_prok/) and Prokatyotic Genome Annotation (Prokka) version 1.12b (Seemann, 2014) respectively. Open Reading Frames (ORFs) were predicted by Rapid Annotation using Subsystem Technology (RAST) server (Aziz et al., 2008) online and Glimmer 3.2 (Delcher et al., 1999).
The complete genome sequence of B. velezensis SRCM102755 was composed of single circular chromosome and is 4,044,843 bp with a G + C content of 46.34%. Based on result of gene prediction, genome of SRCM102755 contains 3,856 CDSs, 86 tRNA, 27 rRNA. We also categorized 3,366 functional protein based on clusters of orthologous genes (COG) designation. The genome of SRCM102755 contains MK-7 synthesis gene cluster comprising nine genes designated as menA (1,4-Dihydroxy-2- naphthoate octaprenyl transferase), menB (1,4-Dihydroxy-2- naphthoate synthase), menC (o-Succinyl benzoic acid synthase), menD (2-Succinyl-6-hydroxy-2,4-cycloheaxdiene-1-carboxylic acid synthase), menE (o-succinyl benzoic acid-CoA synthase), menH (2-Succinyl-6-hydroxy-2,4-cycloheaxdiene-1-carboxylic acid synthase), menG (Demethylmenaquinone methyltransferase), hepS (Heptaprenyl pyrophosphate synthases I) and hepT (Heptaprenyl pyrophosphate synthases II), encoding the head structure biosynthesis enzymes related to the MK-7 biosynthesis starting from isochorismate (Xu et al., 2017).
Nucleotide sequence accession number
The complete genome sequence of Bacillus velezensis SRCM102755 was deposited at NCBI under the GenBank accession no. CP028204.
적 요
Bacillus velezensis SRCM102755는 된장에서 분리하였으 며, menaquinone-7 고함량 청국장을 개발하기 위한 스타터로 선별되었다. B. velezensis SRCM102755 균주 유전체의 염기 서열은 Pacific Biosciences (PacBio) single-molecule real-time (SMRT) RS II platform으로 분석하였으며, 4,044,843 bp 크기 로 GC 비율은 46.34%의 환형 구조를 가지고 있는 것으로 확인 되었고, 유전체 이외의 플라스미드는 존재하지 않았으며, 유전 체는 3,856개의 단백질 코딩 유전자와 113개의 RNA (27 rRNA, 86 tRNA)로 구성되어 있음을 확인하였다. Menaquinone-7 생 합성과 관련된 유전자는 최소 9개 이상 포함되어 있는 것으로 나타났다.
Acknowledgments
This study was supported by the Ministry of Trade, Industry
& Energy (MOTIE), Korea Institute for Advancement of Technology (KIAT) and Establishment of Infrastructure for Industrialization of Korean Useful Microbes (R0004073).
References
