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Biomedical subjects

J T Singer

Publications and source records attributed to J T Singer.

25 records · Page 2Linked to original sources
PubMed1985

Identification of the Escherichia coli deoR and cytR gene products.

The protein products encoded by the Escherichia coli deoR and cytR structural genes have been identified based on results obtained from E. coli maxicells harboring (i) recombinant plasmids carrying wild-type deoR and cytR genes, (ii) deletion derivatives of the deoR+ and cytR+ plasmids, (iii) plasmids containing site-specific mutations in the deoR and cytR structural genes, and (iv) plasmids which have transposon Tn1000 inserted into the deoR and cytR structural genes. Analysis of the protein profiles obtained from all the maxicell experiments demonstrated that the deoR gene encodes a protein with a subunit molecular weight of 30,500 and that the product of the cytR gene is a protein with a subunit molecular weight of 37,000.

Base Sequence↗
PubMed1984

Studies on deo operon regulation in Escherichia coli: cloning and expression of the deoR structural gene.

Recombinant plasmid pBR322 derivatives containing the Escherichia coli deoR structural gene (coding for one repressor of the deo operon) and a mutant allele of the cmlA gene (chromosomally encoded chloramphenicol resistance) have been constructed and the positions of these genes on a 6.3-kb EcoRI fragment have been determined. Transformation of an E. coli deoR single mutant with any of the deoR+ plasmids resulted in complementation of the chromosomal deoR mutation. More importantly, however, transformation of a deoR cytR double mutant with the deoR+ plasmids also resulted in complete repression of Deo enzyme synthesis. Based on these data, we conclude that transcription of the deo operon initiating from both the cAMP/CRP-independent promoter-operator site, PO1, and the cAMP/CRP-dependent promoter-operator site, PO2, is negatively controlled by the deoR-encoded repressor, whereas the cytR-encoded repressor regulates deo operon expression only from the cAMP/CRP-dependent promoter-operator site, PO2.

Bacterial Proteins↗
PubMed1984

Insertional specificity of transposon Tn5 in Acinetobacter sp.

Suicide plasmid pJB4JI, containing transposon Tn5 and phage Mu, was introduced from Escherichia coli 1830 into Acinetobacter sp. strain HO1-N and Acinetobacter calcoaceticus BD413. Kanamycin-resistant (Kmr) exconjugants of HO1-N and BD413, isolated on complex medium, were screened for auxotrophic requirements. Over 10,000 Kmr clones were examined, but no auxotrophs were detected. Several Kmr exconjugants of BD413 and HO1-N, obtained from independent matings, were chosen for further study. All Tn5-containing strains exhibited kanamycin phosphotransferase activity. Kmr strains lacked plasmid DNA as determined by three plasmid screening procedures, and the Kmr phenotype was not transferable by conjugal matings to kanamycin-sensitive BD413, HO1-N, or E. coli HB101. The chromosomal location of Tn5 insertions in independently isolated Kmr exconjugants of BD413 and HO1-N was characterized by restriction endonuclease mapping and hybridization studies. Results obtained from Southern hybridization studies were consistent with a single Tn5-specific insertion site in HO1-N and two such sites in BD413. Phage Mu sequences were not detected in Tn5-containing Acinetobacter sp.

Acinetobacter↗
PubMedDate not supplied

Long-term expression of a fluorescent reporter gene via direct injection of plasmid vector into mouse skeletal muscle: comparison of human creatine kinase and CMV promoter expression levels in vivo.

Expression of a fluorescent reporter gene has been studied using two alternate promoters to transcribe the green fluorescent protein (gfp) from Aequorea victoria. The human cytomegalovirus (CMV) enhancer/ promoter or the human muscle-specific creatine kinase promoter (CKM) were inserted along with the gfp cDNA into a plasmid expression vector based on a modified adeno-associated virus genome. Naked plasmid DNA was injected into the hamstring muscle of mdx mice and gfp gene expression determined from frozen muscle sections taken at 4, 14, and 42 days postinjection. Fluorescence patterns obtained by photomicroscopy and quantitative fluorescence measurements indicated a near-linear increase in the accumulation of the gfp in skeletal muscle during the length of the study, with gfp expression at 42 days being roughly four times the values obtained at 4 days. The levels of expression of gfp from the CKM construct were consistantly higher than for the CMV construct. The CKM promoter/expression vector combination demonstrates significant potential for simple, direct delivery and long-term, high-level expression of genes in skeletal muscle.

Animals↗
PubMedDate not supplied

Molecular characterization of Salmonella enteritidis isolates from Maine poultry and poultry farm environments.

Eighty-six Salmonella enteritidis isolates obtained during a surveillance program of poultry farms in Maine were subjected to phage-typing, plasmid profiling and fingerprinting, outer-membrane polypeptide analysis, and antimicrobial sensitivity testing. Isolates were obtained from a variety of sources, including poultry-farm environmental samples, chicken organ samples, human stool samples, cat feces, and live-trapped rats and mice. These isolates were compared with 21 S. enteritidis isolates originating outside of Maine. Phage types isolated in Maine included 13a (60%); 14b (29%); 23 (5%); 8 (2%); and 2 (2%). All S. enteritidis isolates from Maine carried plasmid DNA, and 97% of these isolates carried a 40.3-megadalton plasmid alone (6%) or in conjunction with several smaller plasmids (91%). All 52 phage-type 13a isolates harbored 40.3- and 3.0-megadalton plasmids. All 25 phage-type 14b isolates carried 3.3- and 1.3-megadalton plasmids, and 22 isolates also carried the 40.3-megadalton plasmid. All isolates displayed highly similar outer-membrane polypeptide profiles and were sensitive to a variety of antimicrobials commonly used against gram-negative organisms. The above data suggest that phage type and plasmid content may be related in the cases of phage-type 13a and 14b isolates, and that traditional plasmid-borne antimicrobial resistance determinants were not present in Maine isolates. Results also indicate that phage-typing can be a valuable epizootiological tool for monitoring the potential spread of these strains throughout the Northeast.

Animals↗
PubMedDate not supplied

Wood-drying condensate from Eastern white pine induced cytotoxicity and genotoxicity in vitro.

Eastern white pine is one of the most important commercial species of wood in the Northeast. Condensates extracted from this wood were tested to detect potential cytotoxicity and genotoxicity in Chinese Hamster Ovary (CHO) cells in the absence of S-9 activation. Cytotoxicity was measured by the Trypan blue exclusion assay, mitotic index (MI) and proliferative rate index (PRI). Genotoxicity was measured by the chromosome aberration (CA) assay and sister chromatid exchange (SCE) analysis. Both cytotoxic and genotoxic effects were observed. Laboratory-generated Eastern white pine condensate reduced the viability of CHO cells. The number of viable cells was roughly inversely proportional to dosage over a range of 91 percent to 58 percent survival in treated groups as compared to 2.4 x 10(5) viable cells (100 percent) in the control. The mitotic index (MI) data also showed an inverse correlation with dosage. The highest scorable dose limited by toxicity was determined to be 1 ml of Eastern white pine condensate in a total of 10 ml of medium. Lastly, a dose response curve was observed using the CA assay and also with the SCE analysis. The present findings support results obtained from Ames testing of Eastern white pine condensate and also corroborate results derived from human peripheral-blood lymphocytes.

Animals↗