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PubMed1972

Identification and genetic characterization of mutants of bacteriophage T4 defective in the ability to induce exonuclease A.

A mutant of bacteriophage T4 unable to induce exonuclease A has been isolated. The mutation responsible for this defect maps between genes 39 and 56, in a region of the chromosome devoid of other known markers. Four deletion mutants lacking part of the genome located between genes 39 and 56 also fail to induce exonuclease A. The ability of all of these mutants to replicate suggests that exonuclease A is not essential for replication of phage T4.

Cell-Free System↗

PubMed2002

The molecular genetic characterization of the 'Bobwhite' bread wheat family using AFLPs and the effect of the T1BL.1RS translocation.

Bobwhite is a generic name that refers to all sister lines derived from the cross CM 33203 with the pedigree Aurora//Kalyan/Bluebird/3/Woodpecker made by the CIMMYT bread wheat program in the early 1970s. Individual sister lines can be distinguished by their unique selection history. One of the parents, Aurora, contains the T1BL.1RS translocation from rye, and approximately 85% of the sister lines have inherited the translocation. The sister lines demonstrate great variability for agronomic traits such as maturity, height, grain color, reaction to leaf rust, stem rust, yellow rust, septoria leaf blotch and powdery mildew. Certain groups of sister lines derived from particular F(1) plants can be distinguished by their phenotype. One hundred and one Bobwhite sister lines were fingerprinted using four AFLP enzyme/primer combinations. Following multivariate analysis, two main and very distinct clusters were found, which reflected the presence or absence of the T1BL.1RS translocation. Within these clusters, lines clustered together, for the most part, with other sister lines sharing a common selection history. Removal of the AFLP markers that were correlated with the presence or absence of the translocation caused lines to cluster based on pedigree alone. Therefore, the presence of translocations in wheat could bias genetic diversity studies using unmapped markers such as AFLPs that are located on the translocated segment(s), with the result that the resulting clusters will not reflect the true degree of relatedness.

Journal Article↗

PubMed1976

Isolation and genetic characterization of the nitA mutants of Escherichia coli affecting the termination factor rho.

Taking advantage of the Spi (sensitivity to P2 interference) phenomenon, bacterial mutants seemingly resistant to phage lambdasusNnin5, but sensitive to phage lambdaspi, were isolated from a strain of E. coli K12 carrying no nonsense suppressor and lysogenic for P2. A class of these mutants, designated nitA (N-independent transcription), is described here. Upon infection of the nitA mutants with a trp transducing phage lambdasusN7N53ptrp46 which carries the E. coli trpE and D genes in the CIII-att region of the lambda genome, formation of anthranilate synthetase (ASase, a complex protein of trp E and D gene products) was clearly demonstrated. In contrast, no ASase formation was observed in the parent nitA+ strain under the same conditions. The synthesis is subject to "turn off" control, and is completely repressed by the CI repressor of phage lambda. The nitA cells lysogenic for lambdaCI857susN7N53 are killed by thermal induction much more efficiently than the parent cells lysogenic for the same phage. The nitA mutants support the growth of lambdasusN7N53byp much better than the parent. These results suggest that the nitA mutation permits the early leftward and rightward transcription of the lambda genome in the absence of the N gene product. On the E. coli genetic map, nitA is located between ilv and metE, nearer to ilv. The mutant allele is recessive to the wild-type allele. The present evidence, together with results of biochemical investigations to be reported, suggests that nitA is a gene specifying the transcription termination factor rho.

Chromosome Mapping↗

PubMed1998

Biochemical and genetic characterization of the DNA ligase encoded by Saccharomyces cerevisiae open reading frame YOR005c, a homolog of mammalian DNA ligase IV.

Here we demonstrate that the Saccharomyces cerevisiae DNA ligase activity, which we previously designated DNA ligase II, is encoded by the genomic DNA sequence YOR005c. Based on its homology with mammalian LIG4, this yeast gene has been named DNL4 and the enzyme activity renamed Dnl4. In agreement with others, we find that DNL4 is not required for vegetative growth but is involved in the repair of DNA double-strand breaks by non-homologous end joining. In contrast to a previous report, we find that a dnl4 null mutation has no effect on sporulation efficiency, indicating that Dnl4 is not required for proper meiotic chromosome behavior or subsequent ascosporogenesis in yeast. Disruption of the DNL4 gene in one strain, M1-2B, results in temperature-sensitive vegetative growth. At the restrictive temperature, mutant cells progressively lose viability and accumulate small, nucleated and non-dividing daughter cells which remain attached to the mother cell. This novel temperature-sensitive phenotype is complemented by retransformation with a plasmid-borne DNL4 gene. Thus, we conclude that the abnormal growth of the dnl4 mutant strain is a synthetic phenotype resulting from Dnl4 deficiency in combination with undetermined genetic factors in the M1-2B strain background.

Animals↗

PubMed1997

Genetic characterization of the pdu operon: use of 1,2-propanediol in Salmonella typhimurium.

Salmonella typhimurium is able to catabolize 1,2-propanediol for use as the sole carbon and energy source; the first enzyme of this pathway requires the cofactor adenosyl cobalamin (Ado-B12). Surprisingly, Salmonella can use propanediol as the sole carbon source only in the presence of oxygen but can synthesize Ado-B12 only anaerobically. To understand this situation, we have studied the pdu operon, which encodes proteins for propanediol degradation. A set of pdu mutants defective in aerobic degradation of propanediol (with exogenous vitamin B12) defines four distinct complementation groups. Mutations in two of these groups (pduC and pduD) eliminate propanediol dehydratase activity. Based on mutant phenotypes, a third complementation group (pduG) appears to encode a cobalamin adenosyl transferase activity. No function has been assigned to the pduJ complementation group. Propionaldehyde dehydrogenase activity is eliminated by mutations in any of the four identified complementation groups, suggesting that this activity may require a complex of proteins encoded by the operon. None of the mutations analyzed affects either of the first two genes of the operon (pduA and pduB), which were identified by DNA sequence analysis. Available data suggest that the pdu operon includes enough DNA for about 15 genes and that the four genetically identified genes are the only ones required for aerobic use of propanediol.

Adenosine↗

PubMed1974

Biosynthesis of ubiquinone in Escherichia coli K-12: biochemical and genetic characterization of a mutant unable to convert chorismate into 4-hydroxybenzoate.

A mutant strain of Escherichia coli unable to carry out the first specific reaction of ubiquinone biosynthesis, that is the conversion of chorismate into 4-hydroxybenzoate, has been isolated. The gene concerned maps at about minute 79 on the E. coli chromosome and has been designated ubiC. This gene is probably the structural gene for chorismate lyase since cell extracts from a transductant strain carrying the ubiC437 mutant allele are unable to convert chorismate into 4-hydroxybenzoate and growing cells of the mutant do not form appreciable quantities of ubiquinone unless 4-hydroxybenzoate is added to the growth medium.

Anthranilate Synthase↗

PubMed2003

[Genetic characterization of the non-structural protein NSP4 from epidemic strains of human rotavirus in China].

BACKGROUND: To clarify the features of gene variation among epidemic strains of human rotavirus NSP4 in China. METHODS: SP4 genes from 27 epidemic strains of human rotavirus isolated in different area of China in recent years were amplified with RT-PCR, the resulted cDNAs were cloned and sequenced. The sequences of full length cDNAs were compared with 10 rotavirus NSP4 sequences available in the GenBank using the Clustal x 1.8 TreeView32 and DNA Star softwares. The G serotype of VP7 was analysed by PCR. RESULTS: The homology of the amino acid among the 27 rotavirus strains isolated in China was 81.7%-99.4%. Based on the variation of amino acid sequence, the virus strains can be divided into two groups, represented by Wa and KUN with the homology of 92.0%-99.4% and 92.0%-98.9% within each group, respectively. The diversity between the two groups were 16.6%-21.0%. The Wa group could further be separated into three subgroups, according to the diversity between those strains and the characterization in the highly variable domain. The association between VP7 serotype and NSP4 genotype was not strong. CONCLUSIONS: The NSP4 gene of human rotavirus epidemic strains in China can be divided into Wa and KUN two groups, Wa group is the main group and contains three subgroups possessing characteristic amino acid sites. Samples isolated in the same year but not in the same area shared higher homology.

Antigens, Viral↗

PubMedDate not supplied

Genetic characterization of kanamycin resistance in Campylobacter coli.

The restriction map of plasmid pIP1433 from Campylobacter coli BM2509, isolated in France, was constructed and the location of the kanamycin (Km) and tetracycline (Tc) resistance markers determined using probes for aphA-3 and tetO genes, respectively. These DNA probes were used to study a second Km- and Tc-resistant strain C. coli UA696, isolated in Canada. In this strain, the Km resistance determinant was located in the chromosome. Like pIP1433, the Km resistance determinant in C. coli UA696 specified a 3'-aminoglycoside phosphotransferase of type III whereas the tetO gene was located on a non-transmissible plasmid.

Campylobacter↗

PubMed1995

Genetic characterization of forty ionizing radiation-sensitive strains of Deinococcus radiodurans: linkage information from transformation.

Natural transformation was used to help define a collection of ionizing radiation-sensitive strains of Deinococcus radiodurans. Three putative rec mutations were identified, as were three pol alleles. Forty of the ionizing radiation-sensitive strains were placed into 16 linkage groups, and evidence obtained indicates that each linkage group consists of a cluster of mutations not more than 1,000 bp apart. In addition, a new class of D. radiodurans mutant was described that, although radioresistant, appears to recover from ionizing radiation-induced DNA damage slowly relative to other strains of D. radiodurans.

DNA Damage↗

PubMed1982

Isolation and genetic characterization of temperature-sensitive mutants of vaccinia virus WR.

One hundred temperature-sensitive mutants of vaccinia virus WR were isolated from virus that had been mutagenized with 5-bromodeoxyuridine or N-methyl-N'-nitro-N-nitrosoguanidine. A rapid screening procedure based on the ability of vaccinia virus to form plaques under liquid overlay medium was used to identify potential mutants among randomly picked plaque isolates or plaques preselected for their small size after temperature shift-up. The preselection technique resulted in a sixfold increase in the number of successful mutant isolations relative to the number of plaques picked. All of the mutants had efficiencies of plating at 39.5 degrees C relative to that at 33 degrees C of 10(-4) or less, and 33 of 40 produced 10% or less of the amount of virus at the nonpermissive temperature (39.5 degrees C) relative to that at the permissive temperature (33 degrees C). Experiments with the fluorescent DNA binding dye Hoechst 33258 demonstrated that 6 of the 100 mutants failed to form characteristic cytoplasmic DNA factories at 39.5 degrees C. To facilitate the functional grouping of such a large number of mutants, a rapid infectious center assay was developed. Thirty of the mutants were assigned to 16 or 17 complementation-recombination groups by using this assay. Recombination experiments have allowed the construction of a genetic map representing 22 mutants in 12 of these groups.

Bromodeoxyuridine↗

PubMed1988

Genetic control of intrachromosomal recombination in Saccharomyces cerevisiae. I. Isolation and genetic characterization of hyper-recombination mutations.

Eight complementation groups have been defined for recessive mutations conferring an increased mitotic intrachromosomal recombination phenotype (hpr genes) in Saccharomyces cerevisiae. Some of the mutations preferentially increase intrachromosomal gene conversion (hpr4, hpr5 and hpr8) between repeated sequences, some increase loss of a marker between duplicated genes (hpr1 and hpr6), and some increase both types of events (hpr2, hpr3 and hpr7). New alleles of the CDC2 and CDC17 genes were recovered among these mutants. The mutants were also characterized for sensitivity to DNA damaging agents and for mutator activity. Among the more interesting mutants are hpr5, which shows a biased gene conversion in a leu2-112::URA3::leu2-k duplication; and hpr1, which has a much weaker effect on interchromosomal mitotic recombination than on intrachromosomal mitotic recombination. These analyses suggest that gene conversion and reciprocal exchange can be separated mutationally. Further studies are required to show whether different recombination pathways or different outcomes of the same recombination pathway are controlled by the genes identified in this study.

Cell Cycle↗

PubMed1986

Physical and genetic characterization of the genome of Lactobacillus lactis bacteriophage LL-H.

Bacteriophage LL-H is a virulent phage of Lactobacillus lactis LL23. A restriction map of the phage genome was constructed with various restriction endonucleases. This chromosome has a 34-kilobase size and seems to be circularly permuted. We used a bank of LL-H restriction fragments to study the expression of five of the seven main phage particle proteins. Immunoblotting experiments permitted the mapping on the chromosome of several genes coding for phage particle proteins. We also show that the gene of the main capsid protein is expressed from its own promoter in an Escherichia coli strain.

Bacteriolysis↗

PubMed1990

Nucleotide sequence and genetic characterization reveal six essential genes for the LIV-I and LS transport systems of Escherichia coli.

The nucleotide sequence of the genes encoding the high affinity, branched-chain amino acid transport systems LIV-I and LS has been determined. Seven genes are present on a 7568-base pair DNA fragment, six of which participate directly in branched-chain amino acid transport. Two periplasmic amino acid-binding proteins are encoded by the livJ (LIV-BP) and livK (LS-BP) genes. These two proteins confer specificity on the LIV-I and LS transport systems. livK is the first gene in a polycistronic message that includes four genes encoding membrane components, livHMGF. The protein products of the livHMGF genes are shared by the two systems. An analysis of the livH and livM DNA sequences suggests that they encode hydrophobic proteins capable of spanning the membrane several times. The LivG and LivF proteins are less hydrophobic, but are also tightly associated with the membrane. Both LivG and LivF contain the consensus sequence for adenine nucleotide binding observed in many other transport proteins. A deletion strain that does not express any of the liv genes was constructed. This strain was used to show that each of the membrane component genes is required for high affinity leucine transport, including two genes, livM and livF, for which no previous genetic evidence had been obtained.

Amino Acid Sequence↗

PubMed1996

Isolation and genetic characterization of pGKL killer-insensitive mutants (iki) from Saccharomyces cerevisiae.

The linear double stranded DNA plasmid pGKL1 encodes the yeast killer toxin complex (Gunge et al., 1981) of which the killing mechanism is not understood. We isolated and characterized eight mutants in Saccharomyces cerevisiae that were insensitive to both the intracellularly expressed 28-kDa killer subunit and the native killer toxin complex. These mutations (iki1 through iki5) were all recessive, and classified into five complementation groups. The iki2 mutation was mapped to a position near the centromere on chromosome XIII. We developed a novel screening system to isolate the DNA fragments complementing the iki mutations from a Saccharomyces gene library, and isolated three DNA fragments that complement the iki1, iki3, and iki4 mutations, respectively.

Cloning, Molecular↗

PubMed2006

Genetic characterization of a new set of recombinant inbred lines (LGXSM) formed from the inter-cross of SM/J and LG/J inbred mouse strains.

A new set of LGXSM recombinant inbred (RI) strains is presented. The RI strain panel consists of 18 remaining strains of the original 55 founding strains. Strain characterization is based on 506 polymorphic microsatellites and 4,289 single nucleotide polymorphisms (SNPs) distributed across the genome. Average microsatellite inter-marker distance is 4.80+/-4.84 Mb or 2.91+/-3.21 F(2) cM. SNPs are more densely spaced at 0.57+/-1.27 Mb. Ninety-five percent of all microsatellite inter-marker intervals are separated by less than 15.00 Mb or 8.50 F(2) cM, while 95% of the SNPs are less than 0.95 Mb apart. Strains show expected low levels of nonsyntenic association among loci and complete genomic independence. During inbreeding, the RI strains went through strong natural selection on the agouti locus on Chromosome 2, especially when the epistatically interacting tyrosinase locus on Chromosome 7 carried the wild-type allele. The LG/J and SM/J strains differ in a large number of biomedically important traits, and they and their inter-cross progeny have been used in multiple mapping studies. The LGxSM RI strain panel provides a powerful new resource for mapping the genetic bases of complex traits and should prove to be of great biomedical utility in modeling complex human diseases such as obesity and diabetes.

Animals↗

PubMed2003

Isolation and genetic characterization of the Neurospora crassa REV1 and REV7 homologs: evidence for involvement in damage-induced mutagenesis.

In a previous paper, we reported that the Neurospora crassa upr-1 gene is a homolog of the yeast gene REV3, which encodes the catalytic subunit of DNA polymerase zeta (polzeta). Characterization of the upr-1 mutant indicated that the UPR1 protein plays a role in DNA repair and mutagenesis. To help understand the mechanisms of mutagenic DNA repair in the N. crassa more extensively, we identified N. crassa homologs of yeast REV1 and REV7 and obtained mutants ncrev1 or ncrev7, which had similar phenotypes to the upr-1 mutant. Mutant carrying ncrev7 was more sensitive to UV and 4NQO, and slightly sensitive to MMS than the wild-type. The sensitivity to UV and MMS of the ncrev1 mutant was moderately higher than that of the wild-type, but the sensitivity to 4NQO of the mutant was similar to that of the wild-type. In reversion assay using testers with base substitution or frameshift mutation at the ad-3A locus, each of ncrev1 and ncrev7 mutants showed lower induced-mutability than the wild-type. Expression of ncrev1 and ncrev7 was found to be UV-inducible like the case of upr-1. Genetic analyses showed that the ncrev7 was identical to mus-26, which belongs to the upr-1 epistasis group, and that the ncrev1 was a newly identified DNA repair gene and designated as mus-42. Interestingly, all three mutants have a normal CPD photolyase gene, however, they showed a partial photoreactivation defect (PPD) phenotype, not completely defective but inefficient in photoreactivation. These results suggest that N. crassa REV homolog genes function in DNA repair and UV mutagenesis through the bypass of (6-4) photoproducts.

DNA↗

PubMed1978

Isolation and genetic characterization of a mutation affecting ribosomal resistance to cycloheximide in Tetrahymena.

A dominant mutation at a new locus affecting resistance to cycloheximide has been isolated by exploiting a synergistic relationship with a previously known mutation for cycloheximide resistance in Tetrahymena. The new mutation (ChxB) was induced in a line homozygous for ChxA and was recovered from that background by a new technique termed interrupted genomic exclusion. Segregation data from the interrupted genomic exclusion suggest that ChxA and ChxB are separate, linked loci showing 30% recombination. Minimal lethal doses of cycloheximide for the four possible combinations of the wild-type and mutant alleles of these two genes are: wild type 6 microgram/ml, ChxA 125 microgram/ml, ChxB 10 microgram/ml, ChxA-ChxB 175 microgram/ml.

Animals↗

PubMed1996

A genetic characterization of differences in the sensitivity to radiation-induced malformation frequencies in the mouse strains Heiligenberger, C57BI, and Heiligenberger x C57BI.

We studied the frequency of malformations induced in two mouse strains (Heiligenberger, C57B1/6J) by exposure to x-rays 3 h after conception. Whereas there was a high number of malformed fetuses in Heiligenberger mice (mostly gastroschises) on day 19 of pregnancy, C57B1 did not respond to radiation exposure shortly after conception with an increased frequency of malformed fetuses. Cross-breeding of both strains revealed that no statistically significant increase in radiation-induced malformations was obtained in the F1 fetuses when the father was Heiligenberger and the mother C57B1. In the opposite case (Heiligenberger mother, C57B1 father) a small but statistically significant increase was observed.

Abnormalities, Radiation-Induced↗