Genomic structure of the amplified P-wr locus
--Surinder Chopra, Peifen Zhang and Thomas Peterson

We have previously reported that the P-wr allele contains a 5- to 10-fold tandem repetition of P gene sequences (Athma and Peterson, MNL 65:46, 1991). Southern blot analysis demonstrated that, except for a 700 bp fragment at the 3' end of the P-rr gene, all other P-rr gene probes (Lechelt et al., Mol. Gen. Genet. 219:225-234) map within the amplified region of the P-wr allele (Athma, unpublished). To determine the fine structure of the P-wr allele, we have analyzed several genomic lambda clones isolated from a P-wr (W23) genomic library. All the lambda clones were overlapping, and each one contained a complete 13.3 kb unit, and varying lengths of flanking sequence representing the 5' or 3' ends of the 13.3 kb sequence (Figure 1). These results show that P-wr contains a tandem array of at least three units of a 13.3 kb sequence. The P-wr repeat structure is further supported by analysis of two YAC clones of 180 and 200 kbp provided by Keith Edwards (ICI); restriction mapping of the YAC DNAs is consistent with the map predicted by tandem repetition of the 13.3 kbp sequence. We plan to fractionate the YACs by plasmid integration and marker rescue and thereby isolate individual P-wr repeat units for sequence comparisons and functional tests.
 
Figure 1. Restriction map of P-wr locus. Composite restriction map was generated from overlapping P-wr genomic lambda clones. Vertical arrowheads indicate endpoints of lambda clones L1, L2, L3 and L4. A full length 13.3 kb direct repeat and the 3' and 5' ends of the flanking repeats are shown as horizontal arrows. DNA fragment wr61 represents a unique sequence at the 3' end of the P-wr cDNA which replaces the 700 bp fragment 14 present in the P-rr allele. Boxes with the same filling pattern represent similar DNA sequences. Transcriptional units with exons (E1, E2, and E3) and introns (Int1 and Int2) are shown below the genomic map.


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