Physical mapping and complete nucleotide sequence of the denV gene of bacteriophage T4
- 1 December 1984
- journal article
- research article
- Published by American Society for Microbiology in Journal of Virology
- Vol. 52 (3), 846-856
- https://doi.org/10.1128/jvi.52.3.846-856.1984
Abstract
Phage T4 deletion mutants that are folate analog resistant (far) and contain deletions in the region of the T4 genome near denV were isolated previously. One of these mutants (T4farP12) expressed normal denV gene activity; another mutant (T4farP13) was defective in the denV gene. The rII-distal (right) physical endpoints of these deletions defined the limits of the interval in which the rII-proximal (left) endpoint of the denV gene should be located. The deletion endpoints were identified by restriction and Southern hybridization analyses of phage derivatives containing deoxycytidine instead of hydroxymethyldeoxycytidine in their DNA. The results of these analyses localized the rII-proximal (left) end of the denV gene to a region between 62.4 and 64.3 kilobases on the T4 physical map. denV+ phage resulted from marker rescue with 2 of 5 denV- alleles tested, using plasmids containing a 1.8-kilobase fragment from this region or a 179-base-pair terminal fragment derived from it. Sequencing of the 179-base-pair fragment from wild-type DNA showed a 130-base-pair open reading frame with its termination codon at the rII-proximal end. Confirmation that this open reading frame is part of the denV coding sequence was obtained by identifying a TAG amber codon in the homologous DNA derived from a denV amber mutant strain. This mutant strain rescued the denV+ allele from plasmids containing the wild-type sequence. An adjacent overlapping restriction fragment was also cloned, permitting determination of the remaining denV gene sequence. Based on these results, the 3'' end of the coding region of the denV locus was mapped to kilobase position 64.07 on the T4 physical map and the 5'' end was mapped to position 64.48.This publication has 31 references indexed in Scilit:
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