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Biomedical subjects

R Burgess

Publications and source records attributed to R Burgess.

71 records · Page 4Linked to original sources
PubMed1980

A new mutation rpoD800, affecting the sigma subunit of E. coli RNA polymerase is allelic to two other sigma mutants.

We have characterized a new mutation rpoD800 affecting the sigma gene of E. coli. Upon tranfer to high temperature, a strain with the rpoD800 mutation ceases growth within 30 min. We find that this mutation renders sigma about 10-fold more thermolabile than the wild type sigma at 45 degrees C in vitro. We have compared the temperature profile for inactivation of wild type and mutant sigma and find that the mutant inactivates at a temperature about 9 degrees C lower than does the wild type. The chromosomal locus affected by rpoD800 is shown to be allelic to the locus affected by the spontaneous mutants ts285 and alt-1. All three mutations result in altered sigma and in altered growth at high temperature. We argue that the single locus affected is the structural gene for the sigma subunit of E. coli RNA polymerase.

Alleles↗
PubMed1978

Mutation affecting thermostability of sigma subunit of Escherichia coli RNA polymerase lies near the dnaG locus at about 66 min on the E. coli genetic map.

The Escherichia coli strain, ts-rnp5, originally described in 1975 by G. D. Burdick and H. Berger, is shown to possess an RNA polymerase (RNA nucleotidyltransferase) sigma subunit with an activity 4--6 times less thermostable at 45 degrees than sigma from wild-type strains. This defect remains associated with the sigma polypeptide through a variety of purification stages, including renaturation of sigma after its elution from sodium dodecyl sulfate/polyacrylamide gels. The mutation responsible for decreased thermostability of sigma, called rpoD1, cotransduces with dnaG and therefore is located at about 66 min of the E. coli genetic map.

Chromosome Mapping↗
PubMed1977

Cerebral monitoring by compressed spectral array.

Application of a frequency analysis technique, the compressed spectral array (CSA), permits the display of cerebral activity recorded by the electroencephalogram (EEG) over long periods of time in a succinct, graphic manner. This report is a description of a system which is being developed to investigate long-term monitoring of the EEGs of critically ill patients. The computer-based methodology which has been used to implement this system is described, with illustrations of CSA-transformed EEGs.

Adult↗
PubMed1976

Rapid micromethod for the purification of Escherichia coli ribonucleic acid polymerase and the preparation of bacterial extracts active in ribonucleic acid synthesis.

A rapid micromethod is described for the preparation of nucleic acid-free extracts from Escherichia coli that involves precipitation with polyethylene glycol. Extracts can be prepared from growing cells in 75 min by three short, low-speed centrifugations. The extract did not inhibit added purified ribonucleic acid (RNA) polymerase, suggesting that major inhibitors of RNA synthesis had been removed. This extract should be ideal for assessing the properties of mutant RNA polymerases. The rapid chromatography of the extracts with step elution from deoxyribonucleic acid- and diethylaminoethyl-cellulose columns resulted in high yields of substantially pure RNA polymerase. We used this technique to purify 35S-labeled RNA polymerase. This system should find application for the purification of small quantities of other bacterial RNA polymerases that share the general chromatographic properties of E. coli RNA polymerase.

Chemical Precipitation↗
PubMedDate not supplied

[Identification of two new p53 target genes through implementation of the modified chromatin immunoprecipitation method and inverse PCR].

Direct target loci for the transcription factor p53 were identified through the employment of a combination of a modified version of chromosomal immunoprecipitation and inverse PCR. Irradiation of Hela cells to drive DNA damage response was followed by sequential chromosomal immunoprecipitation utilizing antibodies which recognize the large subunit of RNA polymerase II and p53. Inverse PCR with degenerate oligonucleotides specific for the p53 binding site was subsequently performed on immunoprecipitated DNA and fragments containing putative p53 target genes were subcloned and sequenced. Two sequences were identified which contain near-consensus p53 binding sites as well as recognition sites for the core transcriptional machinery including RNA polymerase II and Sp1. Cotransfections of vectors containing these sequences linked to a reporter with p53 expression vectors resulted in stimulation of transcription. Application of the technology described herein may result in the identification of target loci for a wide variety of transcription factors.

Amino Acid Sequence↗
PubMedDate not supplied

[Use of the modified method of chromatin immunoprecipitation for the isolation of actively transcribed loci].

A modified version of the chromosomal immunoprecipitation (ChIP) assay was implemented for discrete isolation and characterization of actively transcribed genes. Specifically, it was demonstrated with the gene II/9-1 of Sciara coprophila as a model locus that significant enhancement in the isolation of actively transcribed versus repressed and inactive genes can be achieved through the ChIP methodology. A combination of solid-phase magnetic bead technology with chromosomal immunoprecipitation using antibodies that recognize the large subunit (c) of RNA polymerase II resulted in efficient isolation of the promoter region of gene II/9-1 exclusively during the amplification stage of larval development, when the gene is actively transcribed. It is postulated that the novel technology described herein can be applied to a wide variety of systems for efficient isolation and in vivo assessment of actively transcribed genes regulated by virtually any given transcription factor.

Animals↗