Physical mapping and complete nucleotide sequence of the denV gene of bacteriophage T4

E. H. Radany, L. Naumovski, J. D. Love, K. A. Gutekunst, D. H. Hall, E. C. Friedberg

Research output: Contribution to journalArticle

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Abstract

Phage T4 deletion mutants that are folate analog resistant (far) and contain deletions in the region of the T4 genome near denV have been isolated previously. We showed that one of these mutants (T4farP12) expressed normal denV gene activity, whereas 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 DNAs. 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 two of five 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.

Original languageEnglish (US)
Pages (from-to)846-856
Number of pages11
JournalJournal of Virology
Volume52
Issue number3
StatePublished - 1984

Fingerprint

Bacteriophage T4
physical chromosome mapping
bacteriophages
Terminator Codon
nucleotide sequences
mutants
endpoints
Base Pairing
stop codon
Genes
genes
Bacteriophages
Open Reading Frames
open reading frames
DNA
plasmids
Plasmids
Alleles
Amber
alleles

ASJC Scopus subject areas

  • Immunology

Cite this

Radany, E. H., Naumovski, L., Love, J. D., Gutekunst, K. A., Hall, D. H., & Friedberg, E. C. (1984). Physical mapping and complete nucleotide sequence of the denV gene of bacteriophage T4. Journal of Virology, 52(3), 846-856.

Physical mapping and complete nucleotide sequence of the denV gene of bacteriophage T4. / Radany, E. H.; Naumovski, L.; Love, J. D.; Gutekunst, K. A.; Hall, D. H.; Friedberg, E. C.

In: Journal of Virology, Vol. 52, No. 3, 1984, p. 846-856.

Research output: Contribution to journalArticle

Radany, EH, Naumovski, L, Love, JD, Gutekunst, KA, Hall, DH & Friedberg, EC 1984, 'Physical mapping and complete nucleotide sequence of the denV gene of bacteriophage T4', Journal of Virology, vol. 52, no. 3, pp. 846-856.
Radany, E. H. ; Naumovski, L. ; Love, J. D. ; Gutekunst, K. A. ; Hall, D. H. ; Friedberg, E. C. / Physical mapping and complete nucleotide sequence of the denV gene of bacteriophage T4. In: Journal of Virology. 1984 ; Vol. 52, No. 3. pp. 846-856.
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AU - Hall, D. H.

AU - Friedberg, E. C.

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N2 - Phage T4 deletion mutants that are folate analog resistant (far) and contain deletions in the region of the T4 genome near denV have been isolated previously. We showed that one of these mutants (T4farP12) expressed normal denV gene activity, whereas 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 DNAs. 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 two of five 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.

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