Number of the records: 1
Comparative analysis of oligonucleotide primers for high-throughput screening of genes encoding adenylation domains of nonribosomal peptide synthetases in actinomycetes
- 1.
SYSNO ASEP 0451178 Document Type J - Journal Article R&D Document Type Journal Article Subsidiary J Článek ve WOS Title Comparative analysis of oligonucleotide primers for high-throughput screening of genes encoding adenylation domains of nonribosomal peptide synthetases in actinomycetes Author(s) Bakal, Tomáš (MBU-M) RID
Goo, K.-S. (JP)
Najmanová, Lucie (MBU-M) RID
Plháčková, Kamila (MBU-M) RID
Kadlčík, Stanislav (MBU-M) RID, ORCID
Ulanová, Dana (MBU-M)Source Title Antonie van Leeuwenhoek International Journal of General and Molecular Microbiology. - : Springer - ISSN 0003-6072
Roč. 108, č. 5 (2015), s. 1267-1274Number of pages 8 s. Language eng - English Country NL - Netherlands Keywords Nonribosomal peptide synthetase ; Adenylation domain ; Actinomycetes Subject RIV EE - Microbiology, Virology R&D Projects ED1.1.00/02.0109 GA MŠMT - Ministry of Education, Youth and Sports (MEYS) Institutional support MBU-M - RVO:61388971 UT WOS 000362881400024 DOI 10.1007/s10482-015-0557-5 Annotation In the biosynthesis of diverse natural bioactive products the adenylation domains (ADs) of nonribosomal peptide synthetases select specific precursors from the cellular pool and activate them for further incorporation into the scaffold of the final compound. Therefore, the drug discovery programs employing PCR-based screening studies of microbial collections or metagenomic libraries often use AD-coding genes as markers of relevant biosynthetic gene clusters. However, due to significant sequence diversity of ADs, the conventional approach using only one primer pair in a single screening experiment could be insufficient for maximal coverage of AD abundance. In this study, the widely used primer pair A3F/A7R was compared with the newly designed aa194F/aa413R one by 454 pyrosequencing of two sets of actinomycete strains from highly dissimilar environments: subseafloor sediments and forest soil. Individually, none of the primer pairs was able to cover the overall diversity of ADs. However, due to slightly shifted specificity of the primer pairs, the total number and diversity of identified ADs were noticeably extended when both primer pairs were used in a single assay. Additionally, the efficiency of AD detection by different primer combinations was confirmed on the model of Salinispora tropica genomic DNA of known sequence Workplace Institute of Microbiology Contact Eliška Spurná, eliska.spurna@biomed.cas.cz, Tel.: 241 062 231 Year of Publishing 2016
Number of the records: 1