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TOAST - A Tool for the design of targeted gene amplicons for the application of high throughput sequencing in genomic studies of tuberculosis

L Wang(1) M Campos(1) D Ward(1) M Higgins(1) S Campino(1) T Clark(1) J Phelan(1)

1:London School of Hygiene and Tropical Medicine

The targeting of Mycobacterium tuberculosis genes linked to drug resistance and strain-types through amplicon sequencing is a cost-effective approach to profiling infections and personalising clinical management of tuberculosis disease. However, such assays need to be updated with growing knowledge of loci and mutations involved in important phenotypes, including drug resistance and strain-types. Here we present a software tool called TOAST (Tuberculosis Optimized Amplicon Sequencing Tool), which optimises the design of amplicons across platform (e.g., Illumina and Oxford Nanopore Technology) and read length, drug resistance loci, the spoligotyping region; all informed by a growing database of >200K mutations across >100K M. tuberculosis strains. TOAST provides an optimal number and set of primers, across a user-specified amplicon size (based on sequencing platform), coverage, and selected genome-wide loci (e.g., genes). We have validated the outputted primers experimentally, and the tool can be readily modified for the design of amplicons for other important pathogens or species.

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