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Open Access Research article

IS-seq: a novel high throughput survey of in vivo IS6110 transposition in multiple Mycobacterium tuberculosis genomes

Alejandro Reyes12, Andrea Sandoval2, Andrés Cubillos-Ruiz2, Katherine E Varley1, Ivan Hernández-Neuta3, Sofía Samper45, Carlos Martín56, María Jesús García7, Viviana Ritacco8, Lucelly López9, Jaime Robledo1011, María Mercedes Zambrano2, Robi D Mitra1* and Patricia Del Portillo113*

Author affiliations

1 Center for Genome Sciences & Systems Biology, Washington University School of Medicine, St. Louis, MO, 63108, USA

2 Molecular Genetics, Corporación Corpogen, Bogotá, DC, Colombia

3 Molecular Biotechnology, Corporación Corpogen, Bogotá, DC, Colombia

4 Hospital Universitario Miguel Servet. IIS Aragón, Zaragoza, Spain

5 CIBER de Enfermedades Respiratorias (CIBERES), Instituto de Salud Carlos III, Madrid, Spain

6 Departamento de Microbiología, Medicina Preventiva y Salud Pública, Universidad de Zaragoza, Zaragoza, Spain

7 Departamento de Medicina Preventiva, Facultad de Medicina, Universidad Autónoma de Madrid, Madrid, Spain

8 Instituto Nacional de Enfermedades Infecciosas Carlos G Malbrán, Buenos Aires, Argentina

9 Departamento de epidemiología, Universidad de Antioquia, Medellín, Colombia

10 Laboratorio de micobacterias, Corporación para Investigaciones Biológicas y, Universidad Pontificia Bolivariana, Medellín, Colombia

11 Centro Colombiano de Investigación en Tuberculosis (CCITB), Medellín, Colombia

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Citation and License

BMC Genomics 2012, 13:249  doi:10.1186/1471-2164-13-249

Published: 15 June 2012

Abstract

Background

The insertion element IS6110 is one of the main sources of genomic variability in Mycobacterium tuberculosis, the etiological agent of human tuberculosis. Although IS 6110 has been used extensively as an epidemiological marker, the identification of the precise chromosomal insertion sites has been limited by technical challenges. Here, we present IS-seq, a novel method that combines high-throughput sequencing using Illumina technology with efficient combinatorial sample multiplexing to simultaneously probe 519 clinical isolates, identifying almost all the flanking regions of the element in a single experiment.

Results

We identified a total of 6,976 IS6110 flanking regions on the different isolates. When validated using reference strains, the method had 100% specificity and 98% positive predictive value. The insertions mapped to both coding and non-coding regions, and in some cases interrupted genes thought to be essential for virulence or in vitro growth. Strains were classified into families using insertion sites, and high agreement with previous studies was observed.

Conclusions

This high-throughput IS-seq method, which can also be used to map insertions in other organisms, extends previous surveys of in vivo interrupted loci and provides a baseline for probing the consequences of disruptions in M. tuberculosis strains.