Figure 1.

(A) Representation of the 221 VSG expression site. The VSG, pseudo-VSG (Ψ gene), and ESAGs are represented by boxes. ESAGs are numbered according to the nomenclature of Pays et al. (8) for the AnTat 1.3A ES and were determined by analysis of steady-state RNA (7) and complete or partial sequence analysis. The positions of genes in the 221 ES corresponding to ESAGs 2, 4, and 5 from the AnTat 1.3A ES have not been determined. This ES is a 60-kb transcription unit, under control of a single promoter (flag). The recombinant inserts used to form probe mix 4 are depicted as black bars underneath the ES. Numbers refer to recombinant clones pTg221.11, pTg221.14, pTg221.4, and pTg221.8 described in Kooter et al. (7). A detailed description of the probes is given in Materials and Methods. (B) Map of the 3174 transformant of T. brucei variant 221a (36). The single-copy marker gene cassettes (H, hyg; N, neo) are represented by black rectangles flanked by hatched boxes (which correspond to the processing signals). Other symbols are the same as in A. (C) Differences in the genomic integration of the constructs RPhygro and r4 in the rDNA. The top drawing represents the RPhygro transfectant, which contains a hyg cassette (H); the middle drawing shows part of an rDNA array (wild-type); the bottom drawing represents the r4 transfectant, which contains a neo cassette (N). The open box labeled 18S corresponds to the 18S rRNA gene. The constructs are bordered by the restriction enzyme at the integration site (a, AvaI; c, ClaI). In the middle drawing, the black bars represent the homology region used in the constructs. The flags represent transcription initiation (open flag: endogenous rDNA promoter; filled flag: duplicated rDNA promoter).