摘要:

Two hundred thirty-seven independent somatic cell hybrids have been obtained between opossum (Monodelphis domestica) splenocytes, bone marrow cells, or primary fibroblasts, and HPRT-deficient or TK-deficient Chinese hamster, mouse, American mink, or common vole fibroblast lines. Because extreme segregation and fragmentation of marsupial chromosomes commonly occurs in eutherian × marsupial somatic cell hybrids, we developed a rapid primary screening method that enables the identification of primary clones containing a large amount of opossum DNA 20–25 d after fusion. This method, which depends on in situ hybridization of biotin-labeled total opossum DNA on interphase nuclei of hybrid cells fixed on the bottom of microwell plates, was used to screen the 237 hybrid clones; 52 of them had a substantial amount of opossum DNA. G-banding and in situ hybridization of biotin labeled total opossum DNA on metaphase spreads of the clones enabled identification of 17 hybrid clones containing from two to seven intact chromosomes of M. domestica on the background of Chinese hamster or vole chromosomes. The hybrid clones with intact opossum chromosomes are used in a panel constructed for mapping the opossum genome. Initial mapping results from these clones have led to the tentative assignment of GPI and GOT1 to chromosome 1; 6PGD to chromosome 4; LDHA to chromosome 5; LDHB to chromosome 8; and PGK and G6PD to the X chromosome. On the basis of indirect evidence we also tentatively assigned HPRT to the X chromosome and TK to chromosome 5 of M. domestica. These are the first tentative chromosomal assignments by any technique for this speci

ISSN:1424-8581    

DOI:10.1159/000134528

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

Using the methylase Sssl enzyme, we have analyzed the degree of in situ methylation of human sperm pronuclear chromosomes obtained by fertilizing hamster oocytes with human sperm. Untreated (control) sperm chromosome complements showed a higher degree of in situ methylation, compared to sperm complements previously treated with 5-azadeoxycytidine or lymphocyte chromosomes. This indicates that human sperm pronuclear chromosomes have a lower degree of genomic methylation compared to that of other somatic cells. The similarity in the degree of in situ methylation of the euchromatic and heterochromatic regions of chromosomes 1,9, 15, and 16 and the Y chromosome in human sperm does not support the existence of a possible correlation between hypomethylation and heterochromatin decondensation.

ISSN:1424-8581    

DOI:10.1159/000134529

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

The reverse transcriptase inhibitor 3’-azido-deoxythymidine (AZT) has previously been shown to be incorporated into specific regions near the telomeres and centromeres of Chinese hamster ovary cell chromosomes. Our investigation of the effects of AZT on chromosome stability has led to the discovery of a high frequency amplification of telomere-like centromeric DNA. The amplified structures, when analyzed cytogenetically, appear as tandem arrays of tightly clustered blocks of centromeric repeats containing telomeric sequences (TTAGGG)n. There were 5–13 blocks of amplified DNA per structure. These structures form rapidly within one or two cell cycles and can be observed with an incidence as high as 2%. Because the amplification was so rapid, we tested whether the amplification structures could be the result of aberrant overreplication by analyzing BrdU incorporation. Our results indicate that the amplified DNA does not undergo abnormal replication during its formation, but appears to form from existing centromeric regions. We propose a model that involves the excision of multiple centromeric DNA regions from other chromosomes and their relocalization to a new s

ISSN:1424-8581    

DOI:10.1159/000134530

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

The frequency of sperm aneuploidy was investigated by fluorescence in situ hybridization (FISH) in a Hodgkin’s disease patient shortly after he had received chemotherapy and radiotherapy. Sperm karyotyping of the same patient had previously shown multiple structural abnormalities in most spermatozoa immediately after radiotherapy (day 0), whereas most spermatozoa collected 5 wk later (day 38) exhibited normal metaphase divisions (Rousseaux et al., 1993). Variations in the frequency of aneuploidy could not be detected by sperm karyotyping. Multicolor FISH on interphase spermatozoa revealed an increase in the rate of disomy for chromosomes 1, 6, 11, X, and Y at day 0 as well as at day 38. The high frequency of 24, XY (nondisjunction at meiosis I) and 24, XX (nondisjunction at meiosis II) spermatozoa (8.46% and 1.64% at day 0, respectively) from the Hodgkin’s disease patient suggests that both meiosis I and II are affected and that the X chromosome is frequently involved in such malsegregation events. The rate of 46, XY diploidy was also increased in the patient’s sperm, up to 0.62% at day 0. While radiotherapy probably affected the postmeiotic cells (spermatids), the patient’s cancer and/or chemotherapy are the two major factors that could have affected the dividing spermatogonia and/or spermatocytes, resulting in high aneuploid

ISSN:1424-8581    

DOI:10.1159/000134531

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

In eukaryotic cells, protein transport between the endoplasmic reticulum and Golgi compartments is mediated in part by non-clathrin-coated vesicular coat proteins (COP). Seven COP subunits have been recognized, and represent components of a complex known as coatomer. We have previously isolated the cDNA of the human homolog of α-COP, designated HEP-COP and given the official gene symbol COPA. Here we report the genomic organization of COPA, which contains 33 exons ranging in size from 67 to 611 bp. Mapped by PCR and cycle sequencing, all the exon-intron junctions conformed with the GT-AG rule, the 32 introns ranging from about 80 bp to 4 kbp, with the genomic DNA of COPA estimated to span ∼ 37 kb. Southern blot analysis of genomic DNAs of nine eukaryotic species, from human to yeast, revealed identical signals totaling 36 kb each for man and monkey only. Using 5’ RACE and primer extension analysis, the putative transcriptional start site was localized to 466 nucleotides upstream of the translation initiation codon. Comprising a 126-nucleotide 5’ untranscribed genomic sequence and a 466-nucleotide 5’ noncoding cDNA sequence, the 592-nucleotide 5’ CpG island lacked TATA and CAAT boxes but displayed a high G+C content, was enriched for CpG dinucleotides, and contained a potential Spl-binding site, i.e., features compatible with a housekeeping gene. COPA was mapped by fluorescence in situ hybridization to chromosome regio

ISSN:1424-8581    

DOI:10.1159/000134532

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

The chromosome assignment of 115 expressed sequence tags (ESTs) from human skeletal muscle, 101 of which identify unknown human genes, is reported. The ESTs were selected among over 4,000 obtained from systematic sequencing of a skeletal muscle cDNA library containing 3’ portions of the mRNAs. Chromosome assignments were ob tained by PCR amplification of two panels of human × rodent somatic cell hybrids. Analysis of these preliminary data suggests a nonrandom distribution of muscle ESTs in the human chromosome complement. The unexpected occurrence of multiple chromosome localizations for some ESTs is discuss

ISSN:1424-8581    

DOI:10.1159/000134533

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

We report significant heterozygosity for numerous Robertsonian translocations in the southern short-tailed shrew (Blarina carolinensis) in western Tennessee. Eight Robertsonian rearrangements were documented using G-banding techniques that explain the variability in diploid numbers from 46 throughout most of the range of the species to 34–40 in western Tennessee. These fusions resulted in the loss of telomere sequences and were not associated with nucleolar organizer regions. When heterozygocity is considered, the lowest diploid number possibly present would be 30. Four localities with distances of over 180 km apart were sampled, and 80–90% of the collected animals were heterozygous for at least one rearrangement. No putative parental type was found in western Tennessee. Heterozygosity for the same rearrangements was found in these different localities, and no monobrachial fusions were noted. Thus, this is a very wide hybrid zone with rare or absent parental types in the areas sampled or is an evolutionary stage preceding establishment of Robertsonian races. Selective forces, if any, were minimal, as evidenced by the wide area of polymorphism, significant heterozygosity, and the fact that the Robertsonian translocations were in Hardy-Weinberg equilibrium. The origin of such extensive polymorphism in western Tennessee is discussed, especially in light of putative effects of the New Madrid seismic activity. Similarities and differences are noted between the Blαrinα model and the well-documented variation in the European common shrew (Sorex αrαneus) and Mus musculus

ISSN:1424-8581    

DOI:10.1159/000134534

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

The black muntjac (Muntiacus crinifrons) has an unusual karyotype of 2n = 8 in females and 2n = 9 in males. We have studied the evolution of this karyotype by hybridising chromosome-specific paints derived from flow-sorted chromosomes of the Chinese muntjac (M. reevesi, 2n = 46) to chromosomes of the black muntjac. The hybridisation pattern allowed us to infer chromosomal homologies between these two species. Tandem and centromeric fusions, reciprocal translocations, and insertions are involved in the reduction of the diploid number from 2n = 46 to 2n = 8, 9. The painting patterns further show complex chromosomal rearrangements in the male black muntjac which involve more than half the karyotype, including both sex chromosomes. Since early meiosis is reported to be normal without any visible inversion loops of the synaptonemal complex, the observed chromosomal rearrangements would lead to heterosynapsis and, therefore, leave a large fraction of the male black muntjac karyotype balanced between the two sexes.

ISSN:1424-8581    

DOI:10.1159/000134535

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

We have cloned and sequenced the murine homologue of the human EXT1 gene. At the protein level, these genes show almost complete identity as divergence is limited to only 5 amino acid positions that are scattered about the whole sequence. In addition, similarity searches identified a protein from chromosome III of C. elegans that shows significant similarity to the human and murine EXT/Ext genes. Using high resolution backcross mapping, the murine Ext1 was mapped at 26.55 cM between D15Mit143 and D15Mit153 on mouse chromosome 15. Therefore, Ext1 is part of an evolutionarily conserved linkage group including SDC2/Hspgl, TKHR/Trhr, EXT1/Extl, MYC/Myc, and TG/Tgn.

ISSN:1424-8581    

DOI:10.1159/000134536

出版商:S. Karger AG    

年代:1997

数据来源: Karger

摘要:

In routine chromosome harvesting of blood lymphocytes it is well accepted that metaphase spreads are obtained from fixed mitotic cells which burst on the surface of slides during the dropping procedure. For confirmation and clarification, fixed mitotic cells were dropped onto coverslips and observed under an inverted microscope during the evaporation of the fixative. Fixed mitotic cells in the metaphase stage first stick onto the surface of the coverslip without changing their three-dimensional shape and they do not burst. Thereafter, when evaporation of the fixative occurs, they slowly flatten until they are spread. This slow process leads to a stretching of chromosomes which may be a prerequisite for high resolution banding patterns. Confocal laser scanning microscopic measurements of the length, thickness, and width of chromosomes after (i) short term evaporation of the fixative, (ii) evaporation of the fixative under routine harvesting conditions and (iii) a prolonged evaporation, confirmed the stretching of chromosomes. The humidity, the temperature, and the drying time of the fixative influence the dynamic flow of the remaining fixative on the slide. This dynamic flow leads to an intensive wash of the fixed mitotic cells with increasing concentrations of acetic acid which is primarily responsible for the better quality of the metaphase spread.

ISSN:1424-8581    

DOI:10.1159/000134537

出版商:S. Karger AG    

年代:1997

数据来源: Karger