Horizontal gene transfer, in which genetic material is transferred from the genome of one organism to another, has been investigated in microbial species mainly through computational sequence analyses. To address the lack of experimental data, we studied the attempted movement of 246,045 genes from 79 prokaryotic genomes into Escherichia coli and identified genes that consistently fail to transfer. We studied the mechanisms underlying transfer inhibition by placing coding regions from different species under the control of inducible promoters. Our data suggest that toxicity to the host inhibited transfer regardless of the species of origin and that increased gene dosage and associated increased expression may be a predominant cause for transfer failure. While these experimental studies examined transfer solely into E. coli, a computational analysis of gene transfer rates across available bacterial and archaeal genomes supports that the barriers observed in our study are general across the tree of life.

We have developed an automatable procedure for reconstructing the tree of life with branch lengths comparable across all three domains. The tree has its basis in a concatenation of 31 orthologs occurring in 191 species with sequenced genomes. It revealed interdomain discrepancies in taxonomic classification. Systematic detection and subsequent exclusion of products of horizontal gene transfer increased phylogenetic resolution, allowing us to confirm accepted relationships and resolve disputed and preliminary classifications. For example, we place the phylum Acidobacteria as a sister group of delta-Proteobacteria, support a Gram-positive origin of Bacteria, and suggest a thermophilic last universal common ancestor.