Elsevier

Research in Microbiology

Volume 170, Issues 6–7, September–October 2019, Pages 280-287
Research in Microbiology

Original Article
Taxonomic assignment of uncultured prokaryotes with long range PCR targeting the spectinomycin operon

https://doi.org/10.1016/j.resmic.2019.06.005Get rights and content
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Abstract

The taxonomic assignment of uncultured prokaryotes to known taxa is a major challenge in microbial systematics. This relies usually on the phylogenetic analysis of the ribosomal small subunit RNA or a few housekeeping genes. Recent works have disclosed ribosomal proteins as valuable markers for systematics and, due to the boom in complete genome sequencing, their use has become widespread. Yet, in the case of uncultured strains, for which complete genome sequences cannot be easily obtained, sequencing many markers is complicated and time consuming. Taking the advantage of the organization of ribosomal protein coding genes in large gene clusters, we amplified a 32 kb conserved region encompassing the spectinomycin (spc) operon using long range PCR from isolated and from uncultured nodular endophytic Frankia strains. The phylogenetic analysis of the 27 ribosomal protein genes contained in this region provided a robust phylogenetic tree consistent with phylogenies based on larger set of markers, indicating that this subset of ribosomal proteins contains enough phylogenetic signal to address systematic issues. This work shows that using long range PCR could break down the barrier preventing the use of ribosomal proteins as phylogenetic markers when complete genome sequences cannot be easily obtained.

Keywords

Spectinomycin operon
Ribosomal proteins
Long range PCR
Frankia
Prokaryote systematics

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