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Biomedical subjects

B D Young

Publications and source records attributed to B D Young.

At least 109 records · Page 6Linked to original sources
PubMed1987

Molecular analysis of relapse in chronic myeloid leukemia after allogeneic bone marrow transplantation.

Four patients with Philadelphia (Ph') positive chronic myeloid leukemia (CML) were studied before, after, and on relapse following allogeneic bone marrow transplantation (BMT). Southern analysis of DNA from cells collected before and at relapse after BMT was performed in order to investigate the origin of the leukemia at relapse. Using minisatellite probes we showed that the relapse occurred in cells of host origin in all four patients and this was confirmed with a Y chromosome specific probe in two male patients who had a female donor. Furthermore, using two probes for the breakpoint cluster region (bcr) on chromosome 22, we showed that leukemic cells at relapse bore identical rearrangements to those in the disease at time of presentation of each patient. We conclude that relapse in all four patients is due to re-emergence of the original leukemic clone.

Bone Marrow Transplantation↗
PubMed1986

Determination of the DNA content of human chromosomes by flow cytometry.

The mean relative DNA content of each human chromosome was calculated from flow karyotypes of ethidium bromide-stained chromosomes obtained from healthy, normal individuals. These values were found to correlate closely with previously published data obtained by photometric scanning of stained, fixed chromosomes. Calculations of the normal variation in DNA content of each human chromosome indicated that chromosomes 1, 9, 16, and Y (chromosomes with large centric heterochromatic regions) were the most variable, followed by the acrocentrics, 13, 14, 15, 21, and 22. Chromosomes 2, 3, 18, and 19 were also found to vary significantly in DNA content. Chromosomes from a number of subjects with extreme heteromorphisms were flow karyotyped to obtain an estimate of the extent of variation in DNA content of each chromosome. The greatest difference between extreme variants was found for chromosome 1 (which differed by 0.82% of the total genomic DNA), followed by 16 and 9. The largest Y-chromosome variant was 85.9% bigger than the smallest. The precise karyotype analysis produced by flow cytometry resolved many differences between chromosome homologs, including some that cannot be readily distinguished cytogenetically. The implications of these findings for detection of chromosome abnormalities by flow karyotype analysis are discussed.

Chromosome Banding↗
PubMed1986

Rearrangement of the bcr gene in Philadelphia chromosome-negative chronic myeloid leukemia.

We studied the clinical, hematologic, cytogenetic, and molecular biologic features of seven patients with Philadelphia (Ph1) chromosome-negative chronic myeloid leukemia (CML). In five cases the hematologic findings were indistinguishable from those of patients with classical Ph1-positive disease. Myeloid cells were studied by chromosome-banding techniques. One patient had a masked Ph1 chromosome (with translocation t(4;9;22)), one had a deletion involving chromosome 16, and one had a small minority population of 22q- cells without 9q+ but otherwise normal metaphases; metaphases from the other four patients were entirely normal. DNA prepared from the myeloid cells was digested with the restriction enzymes EcoRI, HindIII, BamHI and BglII. Southern analysis using a 0.6-kb fragment of the breakpoint cluster region (bcr) gene showed the presence in each patient's DNA of a germline fragment together with a rearranged fragment or fragments with at least one of the restriction enzymes. We conclude that genomic changes in the bcr gene characteristic of CML can be present in the absence of a Ph1 chromosome.

Adult↗
PubMed1985

Regional mapping of two human immunoglobulin V lambda genes and analysis of the V lambda locus in chronic myeloid leukaemia.

The human immunoglobulin V lambda locus has been studied in relation to chromosomal translocations involving chromosome 22. DNA probes for two V lambda genes which belong to different subgroups and do not cross hybridize, were used to show that both V lambda genes are located on the Philadelphia chromosome in chronic myeloid leukaemia (CML). Both genes map in band 22q11 to a region that is bounded on the distal side by the breakpoints for CML 9:22 translocations and on the proximal side by the breakpoint for an X:22 translocation. We have found no evidence for rearrangements or amplification of either V lambda gene in CML, in either the chronic or acute phases of the disease. In K562 cells which are derived from the pleural effusion of a patient with Ph1-positive CML, there appears to be no rearrangement of the V lambda genes, but they are both amplified about four times. We have estimated that the minimum size for the amplification unit in K562 cells is 186 kb.

Animals↗
PubMed1985

Cloning and sequence analysis of an Ig lambda light chain mRNA expressed in the Burkitt's lymphoma cell line EB4.

A cDNA library was constructed from the mRNA of the Ig lambda producing Burkitt's lymphoma cell line, EB4. Overlapping clones encompassing the coding sequence of the Ig lambda mRNA were isolated and sequenced. The predicted amino acid sequence shows a short hydrophobic leader peptide and a mature polypeptide of 217 residues in which V, J and C regions can be distinguished. The V region belongs to subgroup VI and has greatest homology (80%) with the Amyloid-AR protein. The constant region is the Kern- Oz+ isotype. Probing normal human DNA with the subcloned V lambda coding sequence detects one gene at high stringency and a family of 11 members at low stringency. To date, no restriction enzyme site polymorphisms have been detected. The V lambda VI gene is rearranged on both chromosomes of EB4 and is deleted on both chromosomes in the Burkitt's lymphoma cell line BL2.

Base Sequence↗
PubMed1985

Random association of Epstein-Barr virus genomes with host cell metaphase chromosomes in Burkitt's lymphoma-derived cell lines.

The random association of Epstein-Barr virus DNA with host cell metaphase chromosomes of all sizes in Burkitt's lymphoma-derived cell lines was demonstrated by two substantially different techniques, namely fluorescence-activated chromosome sorting and in situ hybridization. The nature and potential importance of this association are discussed.

Burkitt Lymphoma↗
PubMed1984

The isolation of a human Ig V lambda gene from a recombinant library of chromosome 22 and estimation of its copy number.

We report the first isolation and characterisation of a human Ig V lambda gene. The gene was isolated from a recombinant phage library of human chromosome 22 using a mouse Ig lambda cDNA as probe. DNA sequence analysis predicts a short leader peptide interrupted by an intron of 88 nucleotides, and a mature polypeptide of 96-97 amino acids which shares 61% homology with mouse V lambda I chains. Comparison with the amino acid sequence of known human lambda chains of all six subgroups shows agreement at 22/25 low variance positions. However the maximum homology with human chains is 49%, so we conclude that this sequence represents a new IgV lambda subgroup. The coding region is followed by the conserved heptamer, CACAGTG, and nonamer, ACATAAACC, sequences which have been implicated in V-J segment recombination. This gene has the hallmarks of an active V lambda gene including recently identified transcriptional controlling sequences. Probing genomic DNA with the subcloned V lambda gene detects a family of about 10 cross hybridizing members at low stringency and 2 at high stringency. There is limited polymorphism of the V lambda locus.

Base Sequence↗
PubMed1984

Regional mapping of the human immunoglobulin lambda light chain to the Philadelphia chromosome in chronic myeloid leukaemia.

The lambda light chain immunoglobulin constant region (C lambda) locus was mapped on human chromosome 22. A DNA probe containing part of the C lambda locus was isolated from a human chromosome 22 genomic library, and a series of rodent X human somatic cell hybrids (each of which contained different translocated parts of chromosome 22) were constructed and characterized. The hybridization of the C lambda probe to DNA from these cell hybrids was then studied by Southern blot analysis. The results demonstrates that the C lambda locus is situated very close to the translocation breakpoint on human chromosome 22 which is characteristic of chronic myeloid leukaemia, and at least part if not at all of the locus is situated on the Philadelphia chromosome.

Chromosome Mapping↗
PubMed1984

Chromosome analysis by flow cytometry: a review.

The latest developments in the application of flow cytometry to the analysis and fractionation of metaphase chromosomes is reviewed. This includes the preparative techniques, staining and machine technology used in flow karyotype production. In particular, the application of this technology to the study of chromosomal changes and to the construction of chromosomal specific DNA libraries is reviewed.

Chromosome Mapping↗
PubMed1984

Isolation of a series of HLA class I clones from a human chromosome 6 genomic library.

Human metaphase chromosomes were fractionated by a fluorescent-activated cell sorter (FACS II) and the chromosome 6 fraction was sorted. A genomic library was constructed in lambda gtWES cloning arms using a partial EcoRI digest of the chromosomal DNA. We estimate that at least 60% of our library represents chromosome 6 material, and as 1.2 x 10(5) recombinants were obtained, this indicates that the majority of clonable chromosome 6 sequences are represented. The library was screened with a mouse H-2 class I clone and 18 HLA class I recombinants were isolated from 4 x 10(4) plaques.

Cell Fractionation↗
PubMed1982

Ploidy and progesterone-receptor distribution in flow-sorted deciduomal nuclei.

Nuclei from rat uterine deciduomata were analysed and sorted into different ploidy classes in a fluorescence-activated cell sorter. The highest proportion of 8 n and 16 n nuclei was observed on days 6-7 after decidualization. At this time the progesterone receptor content per unit DNA of the octaploid nuclei was less than half that of the diploid nuclei.

Animals↗
PubMed1982

Construction and characterization of genomic libraries from specific human chromosomes.

Highly purified fractions of human chromosomes 21 and 22 were isolated from a suspension of metaphase chromosomes stained with ethidium bromide by using a fluorescence-activated cell sorter (FACS II). Two recombinant DNA libraries, representing chromosomes 21 and 22, were constructed by complete digestion of DNA from these fractions with EcoRI and insertion into the vector lambda gtWES lambda B. Twenty clones selected at random from the chromosome 22 library hybridized to EcoRI-digested human DNA, and five of these clones hybridized to single bands identical in size to the phage inserts. These five single-copy sequences and a clone coding for an 8S RNA isolated by screening the chromosome 22 library for expressed sequences were characterized in detail. Hybridization of all six clones to a panel of sorted chromosomes and hybrid cell lines confirmed the assignment of the sequences to chromosome 22. The sequences were localized to regions of chromosome 22 by hybridization to translocated chromosomes sorted from a cell line having a balanced translocation t(17;22)(p13;q11) and to hybrid cell lines containing the various portions of another translocation t(X;22)(q13;q112). Five clones reside on the long arm of chromosome 22 between q112 and pter, while one clone and an 18S rRNA gene isolated from the chromosome 22 library reside pter and g112. The construction of chromosome-specific libraries by this method has the advantage of being direct and applicable to nearly all human chromosomes and will be important in molecular analysis of human genetic diseases.

Cell Line↗
PubMed1981

Cloning of a representative genomic library of the human X chromosome after sorting by flow cytometry.

A library of 50,000 recombinants representative of the human X chromosome has been constructed. Human X chromosomes were physically separated using a fluorescence-activated cell sorter. The DNA was purified from the chromosomes, digested to completion with the restriction enzyme EcoRI and cloned into the phage lambda gtWES.lambda B. The X-derived nature of the recombinants was confirmed by hybridization to rodent/human cell line DNA containing only the human X chromosome. Such libraries will be particularly useful for the investigation of genetic diseases such as Duchenne muscular dystrophy, where the basic defect has not been elucidated, and of neoplasia, where several specific chromosomal anomalies, particularly for the leukaemias, have been identified.

Bacteriophage lambda↗
PubMed1981

The calculation of sedimentation coefficients in vertical rotors.

A program for the calculation of sedimentation coefficients of molecules centrifuged in sucrose in vertical rotors has been developed. The program has been tested with both protein and RNA of known sedimentation coefficients. The program can accept any shape of gradient in the range 0-70% sucrose and any temperature in the range of 0-60% C. The program can be used with any vertical rotor for which the dimensions are known.

Catalase↗