title: Multilocus sequence typing of Salmonella strains by high-throughput sequencing of selectively amplified target genes.
abstract: Rapid development of next generation sequencing (NGS) technologies in recent years has made whole genome sequencing of bacterial genomes widely accessible. However, it is often unnecessary or not feasible to sequence the whole genome for most applications of genetic analyses in bacteria. Selectively capturing defined genomic regions followed by NGS analysis could be a promising approach for high-resolution molecular typing of a large set of strains. In this study, we describe a novel and straightforward PCR-based target-capturing method, hairpin-primed multiplex amplification (HPMA), which allows for simultaneous amplification of numerous target genes. To test the feasibility of NGS-based strain typing using HPMA, 20 target gene sequences were simultaneously amplified with barcode tagging in each of 41 Salmonella strains. The amplicons were then pooled and analyzed by 454 pyrosequencing. Analysis of the sequence data, as an extension of multilocus sequence typing (MLST), demonstrated the utility and potential of this novel typing method, MLST-seq, as a high-resolution strain typing method. With the rapidly increasing sequencing capacity of NGS, MLST-seq or its variations using different target enrichment methods can be expected to become a high-resolution typing method in the near future for high-throughput analysis of a large collection of bacterial strains.
authors: Cell and Molecular Biology Program, University of Arkansas, Fayetteville, AR 72701, USA.
authors: Pallavi
authors: P
authors: Singh
authors: Steven L
authors: SL
authors: Foley
authors: Rajesh
authors: R
authors: Nayak
authors: Young Min
authors: YM
authors: Kwon
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics*; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D006801:Humans; D058885:Multilocus Sequence Typing / Q000379:methods*; D016133:Polymerase Chain Reaction; D012475:Salmonella / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D012480:Salmonella Infections / Q000382:microbiology*
chemical_list: D001426:Bacterial Proteins