Gene mapping in the rat by mouse-rat somatic cell hybridization: synteny of the albumin and alpha-fetoprotein genes and assignment to chromosome 14

The methods of somatic cell genetics and molecular hybridization were applied to a panel of mouse X rat hepatocyte hybrids segregating rat chromosomes to assign the rat genes coding for two serum proteins, albumin and alpha-fetoprotein (Alb and Afp). The molecular hybridization of DNAs from different hybrids with cloned DNA probes showed that all the hybrid clones possessing the rat Alb gene and expressing it also retained the rat Afp locus, which is not expressed in these hybrids. So the Alb and Afp genes are syntenic in the rat, as in the mouse. Furthermore, the cytogenetic analysis allowed the assignment of these two loci to rat chromosome 14.     

Assignment of the gene for adenine phosphoribosyltransferase on the genetic map of mouse chromosome 8

Two electrophoretic variants of adenine phosphoribosyltransferase (APRT) were identified in a population of wild mice (Mus musculus bactrianus). Breeding tests demonstrated that the APRT variants are under the control of two alleles at an autosomal locus designatedAprt. We have examined the linkage relationships betweenAprt and the markers of chromosome 8 including esterase-1 and the centromere. The recombination distance between the centromere andAprt is 44 ± 7 cM, and that betweenEs-1 andAprt is 25 ± 2 cM, i.e., the probable order of the markers examined is cen-Es-1-Aprt on chromosome 8.     

Fragile X expression studied by clonal analysis and somatic cell hybridization

The expression of the fragile site on the X chromosome was examined in euploid somatic cell hybrids to determine if a normal genome could complement the abnormal genome and suppress fragile X expression. Fibroblasts from the patient showed the fragile X in 8% to 12% of cells, and fibroblasts from the normal individual showed no fragile X expression. Six fibroblast clones from the patient showed a variable frequency of fragile X expression (from 6.6% to 12%), suggesting that the fragile X is not expressed in a clonal fashion. Three clones from the normal individual did not show the fragile X. Four hybrid clones showed the fragile X in 5.6%, 4.0%, 7.0%, and 4.0% of cells, respectively, indicating that fragile X expression is not completely suppressed by the normal genome.     

Localization of the gene coding for adenine phosphoribosyltransferase on the genetic map of chromosome 8 in the mouse

Polymorphism of electrophoretic mobility of adenine phosphoribosyltransferase (APRT) was found in a population of domestic mice, Mus musculus bactrianus. The Aprt gene was mapped using two markers: plasma esterase 1 coded by the gene Es-1 situated at the distance of 26 morgans from the centromere, and a Robertsonian translocation Rb (8.17) 1 Iem which marks the centromere. The results of linkage analysis permitted to localize the gene Aprt at 51 morgans from the centromere, and 25 morgans distal from the gene Es-1 on the genetic map of chromosome 8. It is found that emotional stress does not alter the recombination rate at chromosome 8, when spermatocytes are at the pachytene stage.     

Synaptophysin: structure of the human gene and assignment to the X chromosome in man and mouse.

Synaptophysin is an integral membrane protein of small synaptic vesicles in brain and endocrine cells. We have determined the structure and organization of the human synaptophysin gene and have established the chromosome localizations in man and mouse. Analysis of a cosmid clone containing the human synaptophysin gene (SYP) revealed seven exons distributed over approximately 20 kb, when compared with the previously published cDNA sequence. The exon-intron boundaries have been identified and do not correlate with functional domains. One intron interrupts the 3' untranslated region. Chromosomal localization of the human and murine genes for synaptophysin established the human SYP locus on the X chromosome in subbands Xp11.22-p11.23 and the mouse synaptophysin gene locus (Syp) on the X chromosome in region A-D. In addition, an Eco0109 RFLP has been identified and used in genetic mapping of the human SYP locus and supports the order TIMP-SYP-DXS14 within a span of approximately 4-7 centimorgans.     

Genetic heterogeneity in Niemann-Pick C disease: a study using somatic cell hybridization and linkage analysis.

The primary molecular defect underlying Niemann-Pick C disease (NPC) is still unknown. A wide spectrum of clinical and biochemical phenotypes has previously been documented. Indication of genetic heterogeneity has recently been provided for one patient. In the present study, somatic cell hybridization experiments were carried out on skin fibroblast cultures from 32 unrelated NPC patients covering the range of known clinical and biochemical phenotypes. The criterion for complementation was the restoration of a normal intracellular fluorescent pattern in polykaryons stained with filipin to document cholesterol distribution. Crosses between the various cell lines revealed a major complementation group comprising 27 unrelated patients and a second minor group comprising 5 patients. Linkage analysis in one multiplex family belonging to the minor complementation group showed that the mutated gene does not map to the 18q11-12 region assigned to the major gene. Patients in the first group spanned the whole spectrum of clinical and cellular phenotypes. No consistent clinical or biochemical phenotypes was associated with the second complementation group. Three of the five group 2 patients, however, presented with a new rare phenotype associated with severe pulmonary involvement leading to death within the first year of life. No biochemical abnormality specific of either group could be demonstrated with regard to tissue lipid storage pattern, intralysosomal cholesterol storage, and regulation of cholesterol homeostasis. Mutations affecting at least two different genes have thus been shown to underlie NPC. The two gene products may function together or sequentially in a common metabolic pathway affecting intracellular cholesterol transport.     

Toward a complete linkage map of the human X chromosome: regional assignment of 16 cloned single-copy DNA sequences employing a panel of somatic cell hybrids.

Closely linked restriction fragment length polymorphisms (RFLPs) are potentially useful as diagnostic markers of genetic defects, and, in principle, RFLPs can be employed to construct a complete linkage map of the human genome. On the X chromosome, linkage studies are particularly rewarding because in man more than 120 X-linked genes are known. Thus, it is probable that each X-specific RFLP will be of use as a genetic marker of one or several X-linked disorders. To facilitate the search for closely linked RFLPs, we have regionally assigned 16 cloned DNA sequences to various portions of the human X chromosome, employing a large panel of somatic cell hybrids. These probes have been used to correlate genetic and physical distances on Xp, and it can be extrapolated from these data that the number and distribution of available Xq sequences will also suffice to span the long arm of the X chromosome.     

Bovine chromosome mapping with the cell hybridization technic. Localization on the x chromosome of glucose-6-phosphate dehydrogenase, phosphoglycerate kinase, alpha-galactosidase A and hypoxanthine phosphoribosyltransferase]

Twelve hamster x cattle (Bos taurus) cell hybrids showed loss of cattle chromosomes, which allows their use for chromosome mapping of cattle. Genes coding for G6PD, alpha-GAL A, PGK, and (indirectly) HGPRT are synteneic and localized on the X chromosome.     

A linkage map of mouse chromosome 8: further definition of homologous linkage relationships between mouse chromosome 8 and human chromosomes 8, 16, and 19.

Using an interspecific cross, a mouse chromosome 8 linkage map spanning 72 cM has been defined by the segregation of restriction fragment length variants. Linkage and genetic distance were established for 10 loci by analysis of 114 meiotic events and indicated the following gene order: (centromere)-Insr-3.5 cM-Plat-26.3 cM-Crryps/Mel/Jund-3.5 cM-Junb/Ucp-10.5 cM-Mt-1-27.2 cM-Acta2-0.9 cM-Aprt. These data provide further definition of mouse chromosome 8 linkage relationships and the relationship between segments of this chromosome and human chromosomes 8, 16, and 19.     

Gene assignment by means of somatic cell hybrids: a new method for the analysis of data

Summary A statistical approach to the interpretation of data from gene assignment with somatic cell hybrids is presented. The observed data are analysed under a variety of hypotheses. The fit to the hypotheses is compared by means of the likelihood obtained under a given hypothesis. Two of these hypotheses are related to fundamental questions: is a gene responsible for the enzyme observation and if so, is that gene located on a specific chromosome or could it change its position and be sometimes on chromosome j and, in another hybrid line, on chromosome k? The other hypotheses concern the assignment of the gene to just one of the chromosomes.To improve the traditional data analysis approach we considered additional information: the uncertainties and possible errors of laboratory methods in all our calculations and the length of the donor chromosomes in connection with one specific hypothesis.This method allows us to account for the reliability of the investigation methods and the nature of the hybrid lines involved. Data can be evaluated at different error probabilities within a realistic range in order to compare and discuss results.     

Interspecific hybridization between human and mouse somatic cells: Enzyme and linkage studies

Human-mouse somatic cell hybrids have been isolated and examined for enzyme and chromosome constitution. The enzymes assayed were lactate dehydrogenase (LDH), isocitrate dehydrogenase (IDH), NADP-dependent malate dehydrogenase (MDH), glucose 6-phosphate dehydrogenase (G6PD), 6-phosphogluconate dehydrogenase (6PGD), phosphoglucomutase (PGM), and several esterases. Coexpression of mouse and human genomes and formation of heteropolymeric enzymes were observed in seven different hybrid populations for the enzymes LDH, IDH, MDH, and G6PD. Evidence predicated on the absence of chromosomal rearrangements is provided for the lack of genetic linkage in the human genome for these four enzymes, as well as for thymidine kinase.Supported in part by NIH Grants GM 09966 and 1-F1-GM-39,399 from the Institute of General Medical Sciences and by NIH Training Grant HD-32.     

Assignment of the gene for adenosine kinase to chromosome 14 in Mus musculus by somatic cell hybridization.

Evidence is presented for the assignment of the gene for adenosine kinase to Mus musculus chromosome 14 by synteny testing and karyotypic analysis of mouse X Chinese hamster somatic cell hybrid clones. ADOK and two enzymes previously mapped to mouse chromosome 14, NP and ES-10, were expressed concordantly in 29 hybrid clones. Chromosome analysis confirmed this assignment. Syntenic evidence is also presented using several clones of a gene transfer system in which the gene for human HPRT had integrated into modified mouse chromosome 14's.     

Assignment of genes to the human X chromosome by the two-dimensional electrophoretic analysis of total cell proteins from rodent-human somatic cell hybrids.

The technique of two-dimensional (2-D) gel electrophoresis was sued to identify five human X-linked gene products in crude cell extracts of mouse-human and Chinese hamster-human somatic cell hybrids. The human origin of these five polypeptides was demonstrated by their comigration with human fibroblast proteins and their failure to comigrate with polypeptides in extracts from the mouse or hamster parental cells. All five polypeptides were present in extracts of rodent-human hybrids that contained a human X chromosome, but were not found in extracts of cells that lacked a human X chromosome. Chromosome analysis of the hybrid clones revealed that the human X chromosome is both necessary and sufficient for the expression of the five polypeptides, designated pX-24, pX-27, pX-37, pX-40, and pX-56. pX-56 can be identified as the human X-linked enzyme glucose-6-phosphate dehydrogenase (G6PD) (E.C.1.1.1.49), while polypeptides pX-24, pX-27, pX-37 and pX-40 have molecular properties unlike those of known human X-linked gene products. pX-24 appears to be a membrane-bound protein that maps to the distal portion of the long arm of the human X chromosome, while pX-27, pX-37, and pX-40 are soluble proteins that map to the proximal long arm or to the short arm of the human X chromosome. 2-D gel electrophoretic analysis of extracts from somatic cell hybrids provides a general method for identifying polypeptides in crude cell extracts coded for by any specific chromosome and can be used to study primary gene products not previously amenable to genetic analysis.     

Gene assignment in the spider monkey (Ateles paniscus chamek--APC): APE-MYH7 to 2q; AR-GLA-F8C to the X chromosome.

Comparative gene assignment between the spider monkey species Ateles paniscus chamek (APC) and man (HSA) showed conserved syntenic associations despite extensive karyotypic rearrangement between species. Two HSA 14q genes were allocated to APC 2q, being syntenic to other HSA 14q and HSA 15q markers previously assigned to APC 2q, and to HSA 12q genes previously assigned to APC 2p. These findings were consistent with A. geoffroyi chromosome painting with human whole-chromosome probes, indicating that the genus Ateles is karyotypically very rearranged. On the other hand, three human X-linked markers were assigned to the Ateles X chromosome, indicating that this chromosome is evolutionary stable.     

Assignment of the growth hormone receptor gene to bovine chromosome 20 using linkage analysis and somatic cell mapping

A polymorphism was identified in the bovine growth hormone receptor ( GHR ) gene by digesting polymerase chain reaction (PCR) products with the restriction enzyme Alul. Two alleles were segregating in cattle of Bos indicus descent, but one allele appears to be fixed in Bos taurus cattle. GHR was localized to bovine chromosome 20 using bovine-rodent hybrid cell lines and linkage analysis.