PubMed Health⌕ Search

Biomedical subjects

C A Pritchard

Publications and source records attributed to C A Pritchard.

11 recordsLinked to original sources
PubMed1989

Segregation of the Huntington disease region of human chromosome 4 in a somatic cell hybrid.

We have developed an X-irradiation:cell fusion procedure that segregates segments of human chromosomes lacking selectable markers and have used this approach to construct somatic cell hybrids retaining fragments of human chromosome 4 as the only human material. To identify hybrids retaining a small chromosomal fragment in the region of the Huntington disease (HD) gene, we used Southern blot analysis to screen 72 hybrid lines for the presence or absence of seven chromosome 4 single-copy loci. These data, combined with in situ hybridization experiments, identified three hybrids of interest. One of these cell lines, C25, stably retains a 10,000- to 20,000-kb fragment of distal 4p in the vicinity of the HD gene, translocated to a hamster chromosome. Field-inversion gel electrophoresis revealed no evidence of rearrangements in the human DNA present in C25. In combination with similar radiation hybrids, C25 is a valuable tool for isolating DNA probes near the HD gene.

Animals↗
PubMed1989

Isolation and field-inversion gel electrophoresis analysis of DNA markers located close to the Huntington disease gene.

A radiation-induced hybrid cell line containing 10-20 million base pairs of DNA derived from the terminal part of human 4p16 in a background of hamster chromosomes has been used to construct a genomic library highly enriched for human sequences located close to the Huntington disease (HD) gene. Recombinant phage containing human inserts were isolated from this library and used as hybridization probes against two other radiation hybrids containing human fragments with chromosomal breaks in 4p16 and against a human-hamster somatic cell hybrid that retains only the 4p15-4pter part of chromosome 4. Of 121 human phage tested, 6 were mapped distal to the HD-linked D4S10 locus. Since the HD gene is located between D4S10 and the 4p telomere, all of these sequences are likely to be closer to HD than D4S10, and any one of them may be a distal flanking marker for the disease locus. Long-range restriction map analysis performed with a field-inversion gel system shows that the six new loci are distributed in different places within 4p16. Although it is not possible to establish an order for the six sequences with the FIGE data, the results demonstrate that the region detected by these probes must span at least 2000 kb of DNA.

Animals↗
PubMed1988

Chromosome fragmentation by chromosome mediated gene transfer.

Chromosome mediated gene transfer (CMGT) can be used to construct human-mouse somatic cell hybrids containing fragments of human chromosomes. We have constructed two panels of hybrid cells by CMGT: the first panel contains fragments of the human X chromosome and the second panel contains fragments of the Y chromosome. The construction of the Y chromosome panel was facilitated by the cotransfer of chromosomes with a selectable plasmid; selection for the plasmid encoded marker enriched for cells which had incorporated chromosome fragments. This preselection was combined with antibody dependent enrichment using the monoclonal antibody 12E7, which reacts with a Y-encoded antigen. Detailed investigation of the two panels revealed extensive rearrangements of the transferred fragments and the preferential retention of sequences from the centromeric region of the donor chromosomes. These results suggest that CMGT will not be particularly useful in the construction of genetic maps, but nevertheless CMGT can be used for enriching for specific regions of the human genome in somatic cell hybrids.

Animals↗
PubMed1987

Isolation of a sequence which maps close to the human sex determining gene.

A sequence mapping close to the human sex determining gene (TDF) has been isolated from a lambda library constructed with DNA derived from a chromosome transfectant hybrid cell line. This sequence is shown to be present in the DNA of X-Y interchange males at a very high frequency and, based on these studies, it is categorised with the sequence defined by the probe, GMGY3, as the closest known Y chromosome derived marker to TDF. In contrast to GMGY3, however, this locus shares no homology with any other human chromosome. Southern blot analysis also reveals specific hybridization to the Y chromosome of other primates. It therefore defines, for the first time, a conserved and Y chromosome unique locus that is near to TDF.

Base Sequence↗
PubMed1987

Investigation of chromosome-mediated gene transfer using the HPRT region of the human X chromosome as a model.

A panel of over 50 hybrid cells containing varying portions of the long arm of the human X chromosome have been obtained by chromosome-mediated gene transfer (CMGT) of human chromosomes to mouse cells deficient in HPRT. This panel is used to investigate the size and integrity of transfected human chromosome fragments and also to examine the effect of including a selectable DNA plasmid in the transfection mix. Chromosomal rearrangements are found to be generated in the chromosome transfer process, and the human X centromeric region is detected in the transfected cells at an unusually high frequency. Extensive lengths of X chromosome DNA are transferred intact, suggesting potential uses of CMGT in cloning large genes and loci for which only the chromosomal map position is known.

Animals↗
PubMed1987

Genetic analysis of the human Y chromosome by chromosome-mediated gene transfer.

Chromosome-mediated gene transfer (CMGT) can be used to segregate fragments of human chromosomes in human-rodent hybrid cells. As with all somatic cell genetics methods, a selection technique is needed to isolate the hybrid cell lines produced by CMGT. Expression of the MIC2 gene product on the cell surface (the 12E7 antigen) provides an endogenous selectable marker for the human Y chromosome. Using chromosome transfer followed by separation of 12E7 antigen-positive cells on the fluorescence-activated cell sorter, a series of cell lines containing segregated fragments of the Y chromosome have been derived. The possibility of using these fragments to derive fine structural mapping data for the Y chromosome is considered in this review.

Animals↗
PubMed1977

Adaptation of museum specimens for use in anatomical teaching aids.

Colour transparencies are prepared of a re-colourized anatomical specimen after placing labels temporarily in position to indicate specific structures. The specimen is also radiographed to show skeletal and soft tissue structures and radio-opaque materials and letters may used to indicate certain parts. Having mounted the specimen it can then be placed in a teaching carrel together with the radiographs and a back-slide projector for viewing the colour transparencies. The student is guided through an examination of the specimen using a script or soundtape. Frequent cross-reference is made between the actual specimen, the colour transparencies on which certain parts of the specimen have been labelled, and also the radiographs. The examination is followed by self-assessment questions relevant to the specimen and these may include identification of structures on a photograph of the specimen and on duplicate radiographs. In the final section of the aid, the answers of the self-assessment questions are given.

Anatomy↗
PubMedDate not supplied

Mapping the limits of the human pseudoautosomal region and a candidate sequence for the male-determining gene.

The human Y chromosome is composed of two different parts: a pseudoautosomal region shared with the X chromosome which is responsible for sex chromosome pairing and a Y-specific part that encodes the sex determining gene. Previously we have shown that the pseudoautosomal gene MIC2 only rarely recombines between the sex chromosomes and, based on the elevated recombination rates in the pseudoautosomal region, we predicted that this gene would lie close to the Y-specific region. In this report we describe a test of this prediction using long-range restriction mapping techniques. We conclude that MIC2 is less than 200 kilobases (kb) away from Y-specific sequences. During these experiments we have identified an HTF island in a position consistent with the proposed location of the human sex determining gene.

Chromosome Mapping↗