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Biomedical subjects

P E Barker

Publications and source records attributed to P E Barker.

At least 19 recordsLinked to original sources
PubMed1993

A novel approach to the production of clinical-grade dextran.

Efficient unit operations for the production and purification of the enzyme dextransucrase and the biosynthesis and fractionation of dextrans have been developed and tested. An alternative process for the production of clinical-grade dextran has been proposed by integrating these processes to achieve improved dextran product yield and the recovery of fructose as a valuable by-product. The enzyme dextransucrase, which synthesizes dextran from sucrose, has been produced either by non-aerated fed-batch or continuous fermentations, purified by continuous ultracentrifugation and ultrafiltration and used in a bioreactor-separator stage for the biosynthesis of dextran. High molecular weight dextrans have been biosynthesized on continuous counter-current chromatographic bioreactor-separator systems. Membrane filtration and size exclusion continuous chromatography were used for the fractionation of the native dextran and production of clinical-grade dextran. The advantages of the proposed process over the conventional process are related to cost reduction due to the elimination of ethanol usage and recovery, improved product quality control and the production and recovery of fructose as a by-product.

Dextrans↗
PubMed1993

Translocation junctions cluster at the distal short arm of chromosome 1 (1p36.1-2) in human neuroblastoma cells.

Human neuroblastoma cells show at high frequency structural changes of the distal short arm of chromosome 1 (1p). The commonly altered region has been identified in previous loss-of-heterozygosity (LOH) studies to involve deletion of 1p36.1-2. These bands are also the site of constitutional alterations in patients with neuroblastoma. In an approach to define the 1p36.1-2 alterations in more detail we here employ four neuroblastoma cell lines to map translocation breaks involving 1p36.1-2 by fluorescence in situ hybridization (FISH). A chromosomal interval flanked by loci DIS96 and DIS98 contained translocation junctions in each of four lines. This analysis identifies in 1p a restricted genomic region as involved in chromosomal rearrangement in different neuroblastomas. The specificity of neuroblastoma translocation junctions at the molecular level implicates this genomic region in tumor development.

Chromosome Banding↗
PubMed1990

Polyacrylamide gel analysis of polypeptides in gene transfer cell lines.

Differential polypeptide expression in gene transfer cell lines of limited genetic complexity was analyzed as a gene mapping strategy. Subcellular fractionation preceding two-dimensional gel electrophoretic analysis simplified protein patterns and revealed subcellular location of differentially expressed polypeptides. As a model system, human MET oncogene polypeptide was identified in gene transfer lines by this approach. Genes encoding five putative human proteins were identified and provisionally assigned to chromosomal region 7q21-31 or to chromosome 1.

Amino Acids↗
PubMed1990

Gene mapping and cystic fibrosis.

The recent molecular cloning of the gene responsible for cystic fibrosis illustrates the importance of human gene mapping to clinical medicine. Identification of a 3bp detection (delta F508) in the majority of cystic fibrosis patients will result in more accurate DNA-based diagnostic testing in cystic fibrosis families, as well as providing clues to the basic molecular defect and its relationship with alterations of anion transport in cystic fibrosis epithelial cells.

Chloride Channels↗
PubMed1990

Genetic complementation in cystic fibrosis pancreatic cells by somatic cell fusion.

The biochemical defect that underlies the genetic disorder cystic fibrosis (CF) has been proposed to involve an altered regulation of epithelial Cl- permeability by agents such as adenosine 3',5'-cyclic monophosphate (cAMP). We report here the successful complementation of this functional defect achieved by using the technique of somatic cell fusion to introduce the normal CF allele into mutant cells. CF epithelial cells were fused with transfectant mouse fibroblasts that contain the normal human gene. The resulting heterokaryons were examined for restoration of cAMP-activated Cl- transport using an optical assay of Cl- permeability. Our results provide direct evidence for the involvement of the protein product of the normal CF allele in modulating epithelial Cl- permeability.

Animals↗
PubMed1989

Insulin receptor overexpression in a human pre-B acute lymphocytic leukemia cell line with a t(1;19) chromosome translocation near the INSR locus.

Human pre-B acute lymphocytic leukemia (ALL) cell line 697 contains a disease-specific chromosomal translocation, t(1;19), near the insulin receptor locus (INSR). Insulin binding, insulin receptor kinase and cell surface immunofluorescence experiments all show that 697 cells express more surface insulin receptors than 207 cells, a line of pre-B ALL cells that lack the t(1;19). Northern blot analysis confirms that 697 cells have increased levels of mRNA for the insulin receptor. Gene dosage and in situ hybridization analysis indicate that only two copies of the INSR locus are present in 697 cells, one on the translocation chromosome and one on the normal chromosome 19. The results described here demonstrate that transcriptional up-regulation of the INSR locus is correlated with the t(1;19) translocation found in human pre-B ALL cell line 697.

Blotting, Northern↗
PubMed1989

The common acute lymphoblastic leukemia antigen gene maps to chromosomal region 3 (q21-q27).

Complementary DNA and genomic clones corresponding to the gene for the common acute lymphoblastic leukemia antigen (CALLA) were used to investigate the genetic structure and location of the CALLA locus. The gene, which encodes a 100-kDa type II transmembrane glycoprotein, appears to be a single copy locus of greater than 45 kb which is not rearranged in malignancies expressing cell surface CALLA. Cell hybrid analysis indicates that the CALLA-related DNA sequences are found on human chromosome 3. In situ hybridization studies reveal the regional location of the CALLA locus to be 3q21-27.

Animals↗
PubMed1988

Primary structure of c-kit: relationship with the CSF-1/PDGF receptor kinase family--oncogenic activation of v-kit involves deletion of extracellular domain and C terminus.

The protein kinase domains of v-kit, the oncogene of the acute transforming feline retrovirus HZ4-FeSV (HZ4-feline sarcoma virus), CSF-1R (macrophage colony stimulating factor receptor) and PDGFR (platelet derived growth factor receptor) display extensive homology. Because of the close structural relationship of v-kit, CSF-1R and PDGFR we predicted that c-kit would encode a protein kinase transmembrane receptor (Besmer et al., 1986a; Yarden et al., 1986). We have now determined the primary structure of murine c-kit from a DNA clone isolated from a brain cDNA library. The nucleotide sequence of the c-kit cDNA predicts a 975 amino acid protein product with a calculated mol. wt of 109.001 kd. It contains an N-terminal signal peptide, a transmembrane domain (residues 519-543) and in the C-terminal half the v-kit homologous sequences (residues 558-925). c-kit therefore contains the features which are characteristic of a transmembrane receptor kinase. Comparison of c-kit, CSF-1R and PDGFR revealed a unique structural relationship of these receptor kinases suggesting a common evolutionary origin. The outer cellular domain of c-kit was shown to be related to the immunoglobulin superfamily. The sites of expression of c-kit in normal tissue predict a function in the brain and in hematopoietic cells. N-terminal sequences which include the extracellular domain and the transmembrane domain as well as 50 amino acids from the C-terminus of c-kit are deleted in v-kit. These structural alterations are likely determinants of the oncogenic activation of v-kit.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗
PubMed1988

Cystic fibrosis DNA markers in Alabama blacks.

DNA samples from unrelated Alabama blacks with no family history of cystic fibrosis (CF) were analyzed with DNA markers linked to cf. Allelic frequencies of genetic markers detected by probes pmetH, pmetD, XV2c, KM19, and pJ3.11 were compared to those of other populations. Allelic frequencies for pJ3.11, XV2c, and KM19 in Alabama blacks and previously studied Caucasian populations were similar. In contrast, the met locus allelic frequencies in Alabama blacks were markedly different from those in Caucasian populations.

Alabama↗
PubMed1987

KRAS2 as a genetic marker for lung tumor susceptibility in inbred mice.

An Eco-RI restriction fragment length polymorphism occurring in a DNA fragment containing the first exon of the murine KRAS2 gene was shown to correlate with the inherited susceptibility of inbred strains of mice to urethan (CAS: 51-79-6)-induced pulmonary adenomas. Eco-RI digestion of murine DNA yielded four KRAS2-specific fragments. Polymorphic variation occurred in the smallest molecular-weight fragment with alleles of either 0.70 or 0.55 kb in size. Genotyping of 14 inbred strains of mice revealed a correlation between KRAS2 Eco-RI polymorphic variation and the differential susceptibility among inbred strains to development of pulmonary adenomas. Strains with a high incidence of pulmonary adenomas, either spontaneously occurring or in response to carcinogen induction, had the 0.55-kb KRAS2 allele whereas adenoma-resistant strains had the 0.70-kb allele. Analysis of a series of recombinant inbred strains (AXB, BXA) that developed from reciprocal crosses between a highly susceptible strain (A/J) and a highly resistant strain (C57BL/6J) revealed a statistically significant threefold difference in lung tumor susceptibility on the basis of KRAS2 genotype. Further analysis of individual F2 mice of a C57BL/6 female X A/J male cross also demonstrated a threefold difference in tumor susceptibility on the basis of KRAS2 allelic variation.

Adenoma↗
PubMed1986

High molecular weight DNA from fixed cytogenetic preparations.

Cell pellets that have been stored in routine clinical cytogenetic fixative were studied for the presence of intact DNA. A method for the isolation of high molecular weight DNA from fixed cytogenetic preparations of human leukocytes, bone marrow, and cell hybrid cultures is presented. DNA preparations from fixed pellets were cleaved with restriction enzymes, transferred to nitrocellulose filters after agarose gel electrophoresis, and hybridized to radiolabeled probes to demonstrate that fixed cell pellets could yield DNA of sufficient quality for Southern blot hybridization analysis. This protocol may be useful for molecular analysis of DNA from fixed cell pellets of patients who are unavailable for additional sampling.

Bone Marrow↗
PubMed1985

Regional location of T cell receptor gene Ti alpha on human chromosome 14.

The chromosomal location of Ti alpha was determined by hybridization of a radiolabeled cDNA for the alpha chain of human T cell receptor with 12 human X mouse cell hybrid DNAs cleaved with BamHI. Seven hybrids contained human Ti alpha, while the remaining five lacked it. Only human chromosome 14 matched the distribution of human Ti alpha signal across the mapping panel. Hybrids segregating a chromosome 14 translocation were used to demonstrate that Ti alpha is in the region 14pter greater than 14q21. Thus, the alpha and beta chain genes that contribute structural components to the Ti moiety of the human T cell receptor lie on different chromosomes. In humans, the immunoglobulin heavy chain locus and Ti alpha are in different regions of chromosome 14, with Ti alpha more proximal and the immunoglobulin heavy chain locus more distal.

Animals↗