Whole-genome fitness analysis in microbes that uses saturating transposon 
mutagenesis combined with massively parallel sequencing (Tn-seq) is providing a 
measure of the contribution of each gene to a given growth condition. With this 
technique, gene fitness profiles and essential genes are discovered by 
simultaneous analyses of whether the absence of each gene product alters the 
growth kinetics of the bacterium. Here we modify the standard Tn-seq procedure 
to simplify and shorten the process by including delivery of the transposon 
through conjugation and liquid culture enrichment of the mutant pool, creating 
transposon liquid enrichment sequencing (TnLE-seq). To illustrate the success of 
these modifications and the robustness of the procedure, analyses of gene 
fitness of two cultures of the strictly anaerobic bacterium Desulfovibrio 
vulgaris Hildenborough were performed, with growth on lactate as the electron 
donor and sulfate as the electron acceptor. These data demonstrate 
reproducibility and provide a base condition for analysis of fitness changes in 
deletion mutants and in various growth conditions. The procedural modifications 
will facilitate the application of this powerful genetic analysis to microbes 
lacking a facile genetic system. Pilot studies produced 2.5×10(5) and 3.4×10(5) 
unique insertion mutants in the anaerobe Desulfovibrio vulgaris Hildenborough 
grown under typical laboratory conditions in rich medium. These analyses 
provided two similar high-resolution maps of gene fitness across the genome, and 
the method was also applied to growth in minimal medium. These results were also 
compared to the coverage obtained with a ca. 13,000-member cataloged transposon 
library constructed by sequencing transposon insertion sites in individual 
mutants.