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Biomedical subjects

B D Young

Publications and source records attributed to B D Young.

At least 91 records · Page 5Linked to original sources
PubMed1990

Direct sequence analysis of the 14q+ and 18q- chromosome junctions in follicular lymphoma.

Although the t(14;18) chromosome translocation has been demonstrated to be a highly consistent feature of follicular lymphomas, the underlying mechanism generating this fusion has remained uncertain. To examine this question further, a polymerase chain reaction strategy has been devised to permit the amplification and direct sequencing of the resultant 14q+ and 18q- reciprocal junctions. Direct sequence analysis of amplified 14q+ junctions established that 7 of 11 tumors contained a bcl-2 (mbr) sequence fused to an immunoglobulin JH region (five were J6 and two were J5). One of these junctions had an unusual configuration with the bcl-2 and JH sequences separated by a recognizable DH region. This finding suggests that at least some of the junctional sequences, previously thought of as N insertions, may be fragments of unrecognized DH regions. It was also possible to amplify and sequence 18q- junctions using a primer based on the DH recombination signal sequences. Several 18q- junctions were shown to consist of DH/bcl-2 (either mbr or mcr) fusions. In two tumors the 14q+ and 18q- junctions were fully sequenced, and it was demonstrated that the bcl-2 sequence was conserved during mbr and mcr translocations. This contrasts with previous analyses that demonstrated either loss or duplication of several bases at the breakpoints in the bcl-2 gene.

Base Sequence↗
PubMed1990

Monosomy 20: a nonrandom finding in childhood acute lymphoblastic leukemia.

We describe four cases of childhood acute lymphoblastic leukemia with monosomy 20 as the sole cytogenetic abnormality. These cases represent 3.4% of cytogenetically abnormal childhood ALL studied in our institute at diagnosis. The patients presented at similar age, ranging from 31 to 36 months. All four patients remain in first remission with survival time being at least 20 months from the time of diagnosis.

Antigens, CD↗
PubMed1990

Rapid isolation of human chromosome-specific DNA probes from a somatic cell hybrid.

A strategy for the rapid isolation from rodent hybrids of human chromosome-specific probes by enzymatic amplification is described. Synthetic oligonucleotide primers based on the consensus Alu sequence were used to amplify inter-Alu sequence from total human genomic DNA and from a somatic cell hybrid, PNTS-1, containing one homolog of chromosome 5 as its only human complement. Direct sequence analysis of the products from human genomic DNA confirmed their inter-Alu structure and provided a novel means for the examination of the 5' end of the Alu consensus. The amplified sequences from the somatic cell hybrid DNA were cloned into a plasmid vector by blunt-end ligation, yielding clones with inserts in the range 300 to 1000 bp. More than 80% of these clones carried inserts that behaved essentially as single-copy human sequences. Hybridization of a selection of these clones to human DNA, hamster DNA, and the original hybrid DNA confirmed that they were derived from chromosome 5. Direct sequence analysis of the vector/insert boundaries in two clones confirmed that inter-Alu sequences had been cloned. This approach has significant advantages over other methods of isolating chromosome-specific probes from hybrid cells, enabling direct separation and cloning of human DNA probes that can be readily used for mapping studies.

Animals↗
PubMed1990

BCR-ABL and BCR proteins: biochemical characterization and localization.

The Philadelphia translocation results in the expression of a family of chimaeric proteins in which a portion of the bcr protein is fused to c-abl protein. Using antibodies which recognize different portions of the bcr gene and abl gene products we have compared the normal bcr products with their chimaeric counterparts. We first conclude that the enhanced kinase activity of the rearranged bcr-abl products (p210 and p190) is recovered almost exclusively from the cytosolic fraction. This methodology was confirmed by the demonstration that in cells transformed by the Abelson murine leukemia virus (A-MuLV) the gag-abl kinase activity was recovered equally from the membrane and cytosolic fractions, in agreement with previous studies. To determine whether the distribution of kinase activity reflected the bulk distribution of the bcr-abl proteins, in vivo labeling followed by subcellular fractionation was performed. Both normal bcr proteins and the p210 bcr-abl protein were recovered from the cytosolic fraction with little detectable amounts present in other fractions. In vivo labeling was also used to demonstrate that both normal bcr products and the p210 bcr-abl had a relatively long half-life. It is concluded that bcr-abl products, like normal bcr products are located in the cytosolic fraction.

Cytosol↗
PubMed1989

Gene mapping by enzymatic amplification from flow-sorted chromosomes.

A new approach to gene mapping which combines enzymatic amplification with high-resolution flow sorting of human chromosomes has been devised. Reliable amplification from as few as 200 chromosomes has been demonstrated. This method, with particular application to mapping the position of chromosomal translocations, has been used to show that the breakpoint for the constitutional translocation t(11;22)(q23;q11) lies proximal to the genes c-ets-1, Thy-1, and T3 delta and distal to the int-2 gene. The mapping was confirmed by Southern analysis to much larger numbers of chromosomes sorted from the same cell line. Control reactions for the bcl-2 gene on chromosome 18 and the C alpha gene of the IGH locus on chromosome 14 demonstrated the discrimination which can be achieved.

Base Sequence↗
PubMed1989

Co-incident N and K ras gene mutations in a case of AML, restricted to differing cell lineages.

Peripheral blood from a patient with acute myeloid leukaemia (AML) of M5 FAB classification, was shown to have mutations to both the N and K ras genes. Leucophoresed blood was separated on a discontinuous Percoll density gradient to provide fractions enriched for different cell lineages. DNA extracted from these fractions was amplified using the polymerase chain reaction (PCR) technique, and hybridized with oligonucleotide probes specific for the single base mutations previously demonstrated. The N-ras mutation was shown to be restricted to the blast and monocytic cell fractions, concordant with the FAB subtype of M5. The K-ras mutation, however, was present in all fractions, suggesting it had occurred in a multi-potential stem cell representing an earlier stage in the generation of the leukaemia, or possibly an incidental background phenomenon.

Adult↗
PubMed1989

Amplification or rearrangement of the beta-human chorionic gonadotrophin (beta-hCG)--human LH gene cluster is not responsible for the ectopic production of beta-hCG by bladder tumour cells.

The beta-subunit of human chorionic gonadotrophin (hCG) is coded on chromosome 19 by the beta-hCG-hLH gene cluster. Genomic DNA has been isolated from bladder tumour cell lines which ectopically express beta-hCG. The beta-hCG-hLH gene cluster was probed for possible rearrangement or amplification and cells karyotyped for chromosome 19 abnormalities. No rearrangement or amplification of the gene cluster and no consistent abnormalities of chromosome 19 were found. The expression of beta-hCG by bladder tumours is therefore likely to be the result of altered gene regulation and not a rearrangement or amplification of this gene cluster.

Cell Line↗
PubMed1988

Extraction of DNA from small sections of frozen tissue with simultaneous histological examination.

Though analysis of small sections of biopsy material by molecular techniques permits increased sensitivity, it also requires accurate histological examination of the tissue in order to reduce sampling error. A technique for the extraction of DNA from small sections of frozen biopsy material with simultaneous histological examination from adjacent sections is described that may enhance the accuracy of characterisation of the tissue, particularly where there is focal variation. The quality of the DNA obtained enables a full range of molecular studies to be carried out.

Biopsy↗
PubMed1988

Identification of two normal bcr gene products in the cytoplasm.

A monoclonal antibody (7C6) has been derived against a synthetic bcr peptide and used to study normal bcr gene products. The expression of a bcr phosphoprotein of 130 kd was demonstrated, in addition to the previously identified bcr phosphoprotein of 160 kd. Sequential immunoprecipitation demonstrated that both p160 and p130 had determinants from two separate regions of the putative bcr translated sequence. The synthesis of bcr products in Philadelphia positive and negative cells was examined by metabolic labelling and it was shown that the rate of synthesis of the p210 bcr-abl product was comparable with that of the normal bcr products. The in vivo phosphorylation of the p160 exceeded that of the p130 and both normal products were unaffected by the increased phosphorylation of the p210 bcr-abl. There was no evidence with the 7C6 antibody of any normal bcr products larger than 160 kilodaltons. Immunofluorescence analysis by conventional and confocal microscopy identified normal bcr products as cytoplasmic proteins with relatively high expression in the myeloid cell line KGl.

Amino Acid Sequence↗
PubMed1987

The potential of family flow karyotyping for the detection of chromosome abnormalities.

Chromosomes from the mother, father, and child of nine families were stained with ethidium bromide and analysed in flow. These flow karyotypes on average resolved separately the homologues of 4.8 of the offspring's chromosomes. A homologue's relative DNA content (calculated from the flow karyotype) was found to be an accurate marker which could be used to trace that chromosome in a family. In this way the parental origin of 74.4% of the offspring's resolved homologues was determined. In the karyotypically normal families studied no chromosome was found in a child which was clearly different from a homologue present in one of the parents. Using parental flow karyotypes to identify familial heteromorphisms, a number of dysmorphic children were studied in an attempt to detect small "de novo" abnormalities. Although no chromosome abnormality was detected in these cases, the usefulness of family studies was illustrated. In one family a large chromosome 4 homologue was found in the child and this was shown to be similar to one found in the father, suggesting an inherited heteromorphism rather than a clinically significant duplication. Flow analysis of the parents of a patient diagnosed cytogenetically as having an interstitial deletion of the X chromosome revealed the abnormality to be a "de novo" 3;X translocation. It is suggested that flow karyotype analysis in families has potential for the detection of chromosome rearrangements at the limits of resolution of conventional cytogenetics.

Chromosome Aberrations↗
PubMed1987

Subtle abnormalities in the short arm of chromosome 11 in acute myeloid leukemia.

A series of four patients with small deletions of the short arm of chromosome #11 is presented. In two of these patients, deletion of 11p was the sole karyotypic abnormality. When compared with similar reported cases an association with FAB type M4 is apparent. Such cases may often be undocumented, because the deletions can be subtle. One patient with erythroleukemia shows an inversion of chromosome #11 involving band 11p15. Because the patients' fetal hemoglobin (HbF) became raised during the course of the disease, it is postulated that the hemoglobin beta chain gene at 11p15 may have been disrupted.

Adolescent↗