METHODS FOR ANALYSIS OF THE C CISTRON OF TEMPERATE PHAGE 16–3 OF RHIZOBIUM MELILOTI

Genetics ◽  
1980 ◽  
Vol 94 (2) ◽  
pp. 265-276
Author(s):  
LÁszló Orosz
Keyword(s):  
Fine Structure ◽  
Rhizobium Meliloti ◽  
Point Mutations ◽  
Cumulative Effect ◽  
Temperate Phage ◽  
Deletion Mutants ◽  
Temperature Sensitive ◽  
Structure Mapping ◽  
Repressor Protein ◽  
Sensitive Allele

ABSTRACT A series of clear mutants of the temperate phage 16-3 of Rhizobium meliloti were isolated that included various point and deletion mutants ofthe Ccistron, coding for the phage repressor. It was observed that recombinant genotypes, such as c+ and ti (temperature-sensitive allele), which form turbid plaques, can be detected quantitatively as lysogenic colonies and scored even at frequencies as low as 10-6. Point mutations, deletions and the autonomy of intracistronic second-site mutations were characterized by this method. Further analysis has shown that each possible pair from three ti mutants gave non-conditional clear recombinants. It was shown that these latter bear the two initial ti mutations, suggesting a cumulative effect of two conditional mutations on the structure of the repressor protein. The double mutants were utilized in fine-structure mapping of the Ccistron.

scholarly journals CORRELATION BETWEEN MAP POSITION AND PHENOTYPE OF CTI MUTANTS IN THE C CISTRON OF RHIZOBIUM MELILOTI PHAGE 16—3

Genetics ◽  
1980 ◽  
Vol 96 (2) ◽  
pp. 321-329
Author(s):  
Brigitta Dudás ◽  
László Orosz
Keyword(s):  
Genetic Map ◽  
Rhizobium Meliloti ◽  
Phenotypic Expression ◽  
Temperate Phage ◽  
Temperature Sensitive ◽  
Repressor Protein ◽  
Phage Strain ◽  
Interallelic Complementation

ABSTRACT Nine temperature-sensitive clear mutations (Cti) in the C cistron (coding for the repressor protein) of Rhizobium meliloti temperate phage 16—3 were characterized according to the inductive temperature, the immunity of cells lysogenic for these mutant phages to superinfection by homoimmune weak virulent mutants, the phenotype of double-ti mutants and interallelic complementation. The results indicate that mutations of similar phenotypic expression are clustered on the genetic map. Furthermore, it seems probable that the C cistron of the original phage 16—3 is identical to that of the independently isolated phage strain 36.

TEMPERATURE-SENSITIVE MUTANTS BLOCKED IN THE FOLDING OR SUBUNIT ASSMBLY OF THE BACTERIOPHAGE P22 TAILSPIKE PROTEIN. I. FINE-STRUCTURE MAPPING

Genetics ◽  
1980 ◽  
Vol 96 (2) ◽  
pp. 331-352 ◽  
Author(s):  
Donna H Smith ◽  
Peter B Berget ◽  
Jonathan King
Keyword(s):  
Fine Structure ◽  
Polypeptide Chain ◽  
Cell Attachment ◽  
Physical Map ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Structure Mapping ◽  
Bacteriophage P22 ◽  
P22 Tailspike ◽  
Ts Mutations

ABSTRACT As part of a study of protein folding, we have constructed a fine-structure map of 9 existing and 29 newly isolated UV- and hydroxylamine-induced temperature-sensitive (ts) mutations in gene 9 of Salmonella bacteriophage P22. Gene 9 specifies the polypeptide chain of the multimeric tail spikes, six of which form the cell attachment organelle of the phage. The 38 ts mutants were mapped against deletion lysogens with endpoints in gene 9. They mapped in 10 of the 15 deletion intervals. Two- and three-factor crosses between mutants within each interval indicated that at least 31 ts sites are represented among the 38 mutants. To determine the distribution of ts sites within the physical map, we identified the protein fragments from infection of su- hosts with 10 gene 9 amber mutants. Their molecular weights, ranging from 13,900 to 55,000 daltons, were combined with the genetic data to yield a composite map of gene 9. The 31 ts sites were distributed through most of the gene, but were most densely clustered in the central third.—None of the ts mutant pairs tested exhibited intragenic complementation. Studies of the defective phenotypes of the ts mutants (Goldenberg and King 1981; Smith and King 1981) revealed that most do not affect the thermostability of the mature protein, but instead prevent the folding or subunit assembly of the mutant chains synthesized at restrictive temperature. Thus, many of thes ts mutations identify sites in the polypeptide chain that are critical for the folding or maturation of the tail-spike protein.

scholarly journals Fine structure mapping and complementation studies of themetDmethionine transport system inSalmonella typhimurium

1992 ◽  
Vol 60 (1) ◽  
pp. 1-6 ◽  
Author(s):  
Carolyn E. Grundy ◽  
P. D. Ayling
Keyword(s):  
Fine Structure ◽  
Transport System ◽  
Binding Protein ◽  
Osmotic Shock ◽  
Point Mutations ◽  
Binding Activity ◽  
Structure Mapping ◽  
Gene Encoding ◽  
Complementation Groups ◽  
Methionine Transport

SummaryA fine structure deletion map of themetDregion of the chromosome ofSalmonella typhimuriumresponsible for a high-affinity methionine transport system has been constructed. Complementation tests involving the introduction ofmetD+DNA contained in a pUC8 vector intometDstrains indicated the presence of four complementation groups in themetDregion. This suggested that the methionine system belongs to the osmotic shock-sensitive class of transport system, and therefore should possess a periplasmic methionine-binding protein and several membrane proteins. But a deletion mutation covering all knownmetDpoint mutations did not affect the level of a methionine binding activity in osmotic shock fluids, suggesting either that the deletion did not extend into the gene encoding the binding protein, or that the binding activity is not associated with themetDsystem. Possible reasons for the failure to isolate mutations in the gene for the binding protein are discussed.

scholarly journals Intragenic and extragenic suppressors of mutations in the heptapeptide repeat domain of Saccharomyces cerevisiae RNA polymerase II.

Genetics ◽  
1989 ◽  
Vol 123 (4) ◽  
pp. 715-724 ◽  
Author(s):  
M L Nonet ◽  
R A Young
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Large Subunit ◽  
Point Mutations ◽  
Cold Sensitivity ◽  
Deletion Mutants ◽  
Temperature Sensitive ◽  
Repeat Domain ◽  
Cold Sensitive ◽  
Extragenic Suppressors

Abstract The largest subunit of RNA polymerase II contains a repeated heptapeptide sequence at its carboxy terminus. Yeast mutants with certain partial deletions of the carboxy-terminal repeat (CTR) domain are temperature-sensitive, cold-sensitive and are inositol auxotrophs. Intragenic and extragenic suppressors of the cold-sensitive phenotype of CTR domain deletion mutants were isolated and studied to investigate the function of this domain. Two types of intragenic suppressing mutations suppress the temperature-sensitivity, cold-sensitivity and inositol auxotrophy of CTR domain deletion mutants. Most intragenic mutations enlarge the repeat domain by duplicating various portions of the repeat coding sequence. Other intragenic suppressing mutations are point mutations in a conserved segment of the large subunit. An extragenic suppressing mutation (SRB2-1) was isolated that strongly suppresses the conditional and auxotrophic phenotypes of CTR domain mutations. The SRB2 gene was isolated and mapped, and an SRB2 partial deletion mutation (srb2 delta 10) was constructed. The srb2 delta 10 mutants are temperature-sensitive, cold-sensitive and are inositol auxotrophs. These phenotypes are characteristic of mutations in genes encoding components of the transcription apparatus. We propose that the SRB2 gene encodes a factor that is involved in RNA synthesis and may interact with the CTR domain of the large subunit of RNA polymerase II.

scholarly journals A COMPARISON OF TWO-POINT, THREE-POINT AND DELETION MAPPING IN THE C CISTRON OF RHIZOBIOPHAGE 16–3, WITH AN EXPLANATION FOR THE RECOMBINATION PATTERN

Genetics ◽  
1980 ◽  
Vol 94 (2) ◽  
pp. 249-263
Author(s):  
LÁszlÓ Orosz ◽  
Katalin RostÁs ◽  
Rollin D Hotchkiss
Keyword(s):  
Fine Structure ◽  
High Frequency ◽  
Rhizobium Meliloti ◽  
General Tendency ◽  
Deletion Mapping ◽  
Structure Mapping ◽  
Mapping Data ◽  
Correct Order ◽  
Point Mapping ◽  
Double Crossovers

ABSTRACT A general tendency for additivity prevailed in recombination frequencies for two-point fine-structure mapping of 14 mutants in the C cistron of Rhizobium meliloti phage 16-3, with little evidence of any marker effect. Intracistronic three-point mapping indicated that double crossovers are rare. Deletion mapping indicated that the two- and three-point mapping data gave the correct order of the mutations. A high frequency (5 to 8%) of c/c+ heterozygotic phage progenies was observed in standard crosses. This pattern implies formation of a relatively long region of heterozygosity. Together with the results of the three-point tests, it suggests certain properties of the branch migration and resolution steps envisioned in current mechanisms of recombination.

scholarly journals Fine structure mapping and properties of mutations suppressing thelonmutation inEscherichia coliK-12 and B strains

1977 ◽  
Vol 30 (3) ◽  
pp. 273-286 ◽  
Author(s):  
Ben F. Johnson
Keyword(s):  
Escherichia Coli ◽  
Fine Structure ◽  
Growth Conditions ◽  
Temperature Sensitive ◽  
Structure Mapping ◽  
E Coli ◽  
Uv Sensitivity ◽  
Division Process ◽  
Increased Sensitivity ◽  
Derivatives Of

SUMMARYMutations,sulAandsulB, that suppress the UV sensitivity conferred by thelonmutation have been isolated and precisely positioned on the linkage map ofEscherichia coli. TheE. coliB strains Bs-3 and Bs-8 have been shown to possesssulAmutations. Also theE. coliK12 strain J6271 that possesses a suppressor of thelonmutation, previously designated assuf, has been shown to be asulAmutation. A series of methylmethane sulphonate resistant derivatives of anE. coliK12lonstrain has been isolated and genetically characterized. In addition tosulAmutations, a second suppressorsulBwas identified and located betweenleuandazigenes on the chromosome. NeithersulAorsulBmutations result in increased sensitivity to the antibiotics ampicillin, rifampicin, or actinomyein D, nor do they have any significant effect upon the overproduction of mucopolysaccharide caused by thelonmutation. Under some growth conditions thesulBmutation causes cells to be temperature sensitive for the cell division process at 42 °C.

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