YAMAT-seq of mature tRNA pools in the bacterium Pseudomonas fluorescens SBW25 and derivatives.

Mature tRNA pools were measured using an adaptation of YAMAT-seq (Shigematsu et al., 2017; doi:10.1093/nar/gkx005 ) and further described in (Ayan et al., 2020; doi:10.7554/eLife.57947) in eight strains dervied from the model bacterium, Pseudomonas fluorescens SBW25. The aim of the experiment was to determine the effect on the mature tRNA pool of (i) removing multi-copy tRNA genes by genetic engineering, and (ii) duplicating tRNA genes during a subsequent, compensatory evolution experiment. We found that (i) results in a reduction in some tRNA isotypes in the mature tRNA pool, and (ii) results in an increase in a compensatory increase in their expression. Overall design: Three replicates of eight strains: SBW25 (wildtype), rWT (reconstructed wildtype, engineering control), W1-1 (wildtype control from evolution experiment), DEGEG (deletion mutant missing a four-gene tRNA locus: gluTTC-glyGCC-gluTTC-glyGCC), DglyGCC (deletion mutant missing one glyGCC gene), M1-1 (evolved from DEGEG; contains ~1 Mb duplication fragment encompassing 7 tRNA genes), M3-1 (evolved from DEGEG; contains ~0.49 Mb duplication fragment encompassing 1 tRNA gene:glyGCC), M5-1 (evolved from DEGEG; contains ~1 Mb duplication fragment encompassing 7 tRNA genes, plus a non-synonymous point mutation in rplC).