A SURVEY OF THE TRYPANOSOMA-BRUCEI-RHODESIENSE GENOME USING SHOTGUN SEQUENCING

A comparison of the efficiency of sequencing random genomic DNA fragme nts versus random cDNAs for the discovery of new genes in African tryp anosomes was undertaken. Trypanosome DNA was sheared to a 1.5-2.5 kb s ize distribution, cloned into a plasmid and the sequences at both ends of 183 cloned fragments determined. Sequences of both kinetoplast and nuclear DNA were identified. New coding regions were discovered for a variety of proteins, including cell division proteins, an RNA-binding protein and a homologue of the Leishmania surface protease GP63. In s ome cases, each end of a fragment was found to contain a different gen e, demonstrating the proximity of those genes and suggesting that the density of genes in the African trypanosome genome is quite high. Repe titive sequence elements found included telomeric hexamer repeats, 76 bp repeats associated with VSG gene expression sites, 177 bp satellite repeats in minichromosomes and the Ingi transposon-like elements. In contrast to cDNA sequencing, no ribosomal protein genes were detected. For the sake of comparison, the sequences of 190 expressed sequence t ags (ESTs) were also determined, and a similar number of new trypanoso mal homologues were found including homologues of another putative sur face protein and a human leucine-rich repeat-containing protein. We co nclude from this analysis and our previous work that sequencing random DNA fragments in African trypanosomes is as efficient for gene discov ery as is sequencing random cDNA clones. (C) 1997 Elsevier Science B.V .