A physical map of 30,000 human genes.

A map of 30,181 human gene-based markers was assembled and integrated with the current genetic map by radiation hybrid mapping. The new gene map contains nearly twice as many genes as the previous release, includes most genes that encode proteins of known function, and is twofold to threefold more accurate than the previous version. A redesigned, more informative and functional World Wide Web site (www.ncbi.nlm.nih.gov/genemap) provides the mapping information and associated data and annotations. This resource constitutes an important infrastructure and tool for the study of complex genetic traits, the positional cloning of disease genes, the cross-referencing of mammalian genomes, and validated human transcribed sequences for large-scale studies of gene expression.

A gene map of the human genome.

The human genome is thought to harbor 50,000 to 100,000 genes, of which about half have been sampled to date in the form of expressed sequence tags. An international consortium was organized to develop and map gene-based sequence tagged site markers on a set of two radiation hybrid panels and a yeast artificial chromosome library. More than 16,000 human genes have been mapped relative to a framework map that contains about 1000 polymorphic genetic markers. The gene map unifies the existing genetic and physical maps with the nucleotide and protein sequence databases in a fashion that should speed the discovery of genes underlying inherited human disease. The integrated resource is available through a site on the World Wide Web at http://www.ncbi.nlm.nih.gov/SCIENCE96/.

Nuclear matrix antigens in azo dye-induced primary rat hepatomas.

Polyvalent antisera, monoclonal antibodies, and immunotransfer methodology have been used to identify and characterize a group of chromosomal protein antigens which appear during azo dye hepatocarcinogenesis. Experiments were designed to probe for the location and placement of antigens in chromatin according to solubility and possible DNA-binding properties. The majority of nuclear antigens were associated with high-speed DNA-containing pellets after ultracentrifugation of chromatin solubilized with denaturing buffers containing 6 M guanidine-HCl:2% sodium dodecyl sulfate, or 2 M NaCl:5 M urea. The addition of 2-mercaptoethanol or dithiothreitol to guanidine-HCl or sodium dodecyl sulfate solutions resulted in solubilization of nearly all antigens from the DNA pellets, suggesting the presence of complexes (protein:protein and/or DNA:protein) cross-linked with sulfhydryl linkages. Preparation of nuclear matrix from the primary hepatomas under several kinds of conditions indicated these antigens to be components of the residual nuclear matrix, envelope, and/or associated structures. Two-dimensional gel analysis showed most antigens to exist in a range of isoelectric forms, suggesting posttranslational modifications. Studies with monoclonal antibodies prepared to these proteins revealed extensive antigenic homology among the members comprising these fractions. Our results document antigenic differences in the nuclear matrix proteins of primary tumors and their normal tissue counterparts.

Characterization of monoclonal antibodies to novikoff hepatoma cytokeratin antigen p39.

Three stable monoclonal antibody-producing mouse hybridoma lines have been developed which produce high-titer, immunoglobulin M antibodies specific for the Novikoff ascites hepatoma (NAH) Mr 39,000 cytokeratin antigen (p39). Immunotransfer assays of cytoskeletal protein-enriched fractions indicated p39 to be present in a range of rat tissues, including colon, breast, lung, and uterus. Two-dimensional gel immunoblots confirmed that immunoreactivity in the latter tissues was for polypeptides with similar isoelectric points to those of NAH p39; however, reactivity in the colon contained a wide range of additional isomeric forms. Immunohistochemical localization studies with these antibodies revealed enrichment of p39 in the simple epithelia or ductular structures of organs containing this antigen. In all sections examined, glandular epithelia were observed to be only weakly immunoreactive. Additional immunoblot and immunocytochemical analyses with the monoclonal antibodies and polyspecific antisera to NAH cytokeratin suggest the human Mr 40,000 cytokeratin to be similar to but not identical to NAH p39.

Social mechanisms in the population genetics of Tay-Sachs and other lethal autosomal recessive diseases: a computer simulation model.

A computer simulation model was developed to study the effects of various feudal social customs on the incidence of lethal autosomal recessive genes. Populations of 500 individuals were created in which each person was characterized by sibship, genotype, and sex. The numbered sibships were sorted into numerical order. Each individual then sought a mate from 6 or 12 sibships on either side of his or her own sibship. The resulting couples were sorted in order of the husband's sibship number, after which the couples produced children in accordance with the prevailing birth rate. This ordering of sibships made the probability of a gene remaining in one locality higher than the probability of it drifting to distant localities. A lethal autosomal recessive gene was introduced into populations at a starting carrier rate of 10%. This fell after 20 generations to 2.54 +/- 0.17% (SEM) which was significantly less than the 5.00% carrier rate predicted by a deterministic model in which random mating occurred in an infinite population. The excessive loss of the lethal gene was caused by the high incidence of consanguineous marriages and by the occurrence of random inequalities in the distribution of the gene. The avoidance of sib marriages raised the final carrier frequency (3.28 +/- 0.19%) as did reproductive compensation (4.33 +/- 0.26%). The combination of the incest taboo and reproductive compensation raised the final carrier rate to 5.05 +/- 0.28%. It is concluded that the social factors which may have operated over a period of many centuries in the past have had a significant effect on the population genetics of lethal autosomal recessive genes.

Cytokeratin and nonhistone protein antigenic changes in rat liver during azo dye but not hepatotoxin feeding.

Chromosomal proteins from rat liver have been assessed immunologically for changes in specific cytokeratins or primary tumor nonhistone proteins during treatment with an azo dye hepatocarcinogen (3'-methyl-4-dimethylaminoazobenzene), (3'-Me-DAB) or the hepatotoxin alpha-naphthyl-iso-thiocyanate (alpha-NIT). Fisher rats were fed laboratory diets supplemented with either agent and sacrificed sequentially at various intervals. Chromosomal proteins from these livers were electrophoresed in the presence of sodium dodecyl sulfate, transferred to nitrocellulose sheets and reacted with rabbit antisera to Novikoff hepatoma cytokeratin or 3'-Me-DAB induced primary hepatoma dehistonized chromatin. Livers from carcinogen-fed animals exhibited distinct, sequential changes in antigenic nonhistone proteins until the immunological specificity characteristic of the hepatoma was achieved concomitant with the induction of neoplasia. No antigenic changes were observed to occur in hepatotoxin-fed animals. The rat carcinoma-specific cytokeratin antigens p39 and p49 in Novikoff hepatoma were observed to appear as early as three weeks after the start of carcinogen feeding and were present maximally in 23 week livers with in situ carcinoma. These cytokeratins were not detected in alpha-NIT-fed animals. Our results support the concept that the carcinogenic process can be related to temporal changes in the expression of cell-specific cytokeratins in addition to nonhistone chromosomal proteins. Furthermore, these data suggest the expression of these antigenic species to not be the direct result of changes in liver structure and cellular composition associated with carcinogen toxicity; rather, neoplastic transformation is apparently required.

An STS-based radiation hybrid map of the human genome.

We have constructed a physical map of the human genome by using a panel of 83 whole genome radiation hybrids (the Stanford G3 panel) in conjunction with 10,478 sequence-tagged sites (STSs) derived from random genomic DNA sequences, previously mapped genetic markers, and expressed sequences. Of these STSs, 5049 are framework markers that fall into 1766 high-confidence bins. An additional 945 STSs are indistinguishable in their map location from one or more of the framework markers. These 5994 mapped STSs have an average spacing of 500 kb. An additional 4484 STSs are positioned with respect to the framework markers. Comparison of the orders of markers on this map with orders derived from independent meiotic and YAC STS-content maps indicates that the error rate in defining high-confidence bins is < 5%. Analysis of 322 random cDNAs indicates that the map covers the vast majority of the human genome. This STS-based radiation hybrid map of the human genome brings us one step closer to the goal of a physical map containing 30,000 unique ordered landmarks with an average marker spacing of 100 kb.