Journal
MICROBIAL GENOMICS
Volume 6, Issue 3, Pages -Publisher
MICROBIOLOGY SOC
DOI: 10.1099/mgen.0.000336
Keywords
multilocus sequence typing (MLST); meticillin-resistant Staphylococcus aureus (MRSA); dual-barcode multiplexing; nanopore; MinION; molecular typing
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Funding
- National Health Research Institutes, Taiwan, ROC [IV-107-PP-07, PH-107-PP-05]
- Ministry of Science and Technology, Taiwan, ROC [MOST 106-2923-B-400-001-MY3]
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Multilocus sequence typing (MLST) is one of the most commonly used methods for studying microbial lineage worldwide. However, the traditional MLST process using Sanger sequencing is time-consuming and expensive. We have designed a workflow that simultaneously sequenced seven full-length housekeeping genes of 96 meticillin-resistant Staphylococcus aureus isolates with dual-barcode multiplexing using just a single flow cell of an Oxford Nanopore Technologies MinION system, and then we performed bioinformatic analysis for strain typing. Fifty-one of the isolates comprising 34 sequence types had been characterized using Sanger sequencing. We demonstrate that the allele assignments obtained by our nanopore workflow (nanoMLST, available at https://github.com/jade-nhri/nanoMLST) were identical to those obtained by Sanger sequencing (359/359, with 100% agreement rate). In addition, we estimate that our multiplex system is able to perform MLST for up to 1000 samples simultaneously; thus, providing a rapid and cost-effective solution for molecular typing.
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