Open AccessnanoMLST: accurate multilocus sequence typing using Oxford Nanopore Technologies MinION with a dual-barcode approach to multiplex large numbers of samples
Author(s) -
Ci-Hong Liou,
Han Chieh Wu,
Yu-Chieh Liao,
TsaiLing Lauderdale,
I-Wen Huang,
Feng-Jui Chen
Publication year - 2020
Publication title -
microbial genomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.476
H-Index - 28
ISSN - 2057-5858
DOI - 10.1099/mgen.0.000336
Subject(s) - minion , multilocus sequence typing , sanger sequencing , nanopore sequencing , housekeeping gene , multiplex , typing , barcode , biology , computational biology , genetics , dna sequencing , computer science , genotype , gene , operating system , gene expression
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.