Abstract
As part of an integrated mapping and sequencing analysis of genomes, we have developed an approach allowing the characterization of large numbers of cDNA library clones with a minimal number of experiments. Three basic elements used in the analysis of cDNA libraries are responsible for the high efficiency of this new approach: (1) high-density library arrays allowing thousands of clones to be screened simultaneously; (2) hybridization fingerprinting techniques to identify clones abundantly expressed in specific tissues (by hybridizations with labeled tissue cDNA pools) and to avoid the repeated selection of identical clones and of clones containing noncoding inserts; and (3) a computerized system for the evaluation of hybridization data. To demonstrate the feasibility of this approach, we hybridized high-density cDNA library arrays of human fetal brain and embryonal Drosophila with radiolabeled cDNA pools derived from whole mouse tissues. Fingerprints of the library arrays were generated, localizing clones containing cDNA sequences from mRNAs expressed at middle to high abundance (>0.1–0.15%) in the respective tissue. Partial sequencing data from a number of clones abundantly expressed in several tissues were generated to demonstrate the value of the approach, especially for the selection of cDNA clones for the analyses of genomes based on expressed sequence tagged sites. Data obtained by the technique described will ultimately be correlated with additional transcriptional and sequence information for the same library clones and with genomic mapping information in a relational database.
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Adams, M., Kelly, J., Gocayne, J., Dubnick, M., Polymeroppoulos, M., Xiao, H., Merril, C., Wu, A., Olde, B., Moreno, R., Kerlavage, A., McCombie, R., and Venter, J.C.: Complementary DNA sequencing: expressed sequence tags and Human Genome Project. Science 252: 1651–1656, 1991.
Adams, M., Dubnick, M., Kerlavage, A., Moreno, R., Kelley, J., Utterback, T., Nagle, J., Fields, C., and Venter, C.: Sequence identification of 2375 human brain genes. Nature 355: 632–634, 1992.
Augenlicht, L., Wahrman, M., Halsey, H., Anderson, L., Taylor, J., and Lipkin, M.: Expression of cloned sequences in biospsies of human colonic tissue and in colonic carcinoma cells induced to differentiate in vitro. Cancer Res 47: 6017–6021, 1987.
Bantle, J., and Hahn, W.: Complexity and characterization of polyadenylated RNA in the mouse brain. Cell, 8: 139–150, 1976.
Brown, N. and Kafatos, F.: Functional cDNA libraries from Drosophila embryos. J Mol Biol 203: 425–437, 1988.
Chomczynski, P. and Sacchi, N.: Single step method of RNA isolation by acid guanidinium-thiocynate-phenol-chloroform extraction. Anal Biochem 162: 156–159, 1987.
Cleveland, D., Lopata, M., MacDonald, R., Cowan, N., Rutter, W., and Kirschner, M.: Number and evolutionary conservation of α-and β-tubulin and cytoplasmic β- and γ-actin. Genes using specific cloned cDNA probes. Cell 20: 95–105, 1980.
Craig, A., Nizetic, D., Hoheisel, J., Zehetner, G., and Lehrach, H.: Ordering of cosmid clones covering the herpes simplex virus type I (HSV-I) genome: a test case for fingerprinting by hybridization. Nucleic Acids Res 18: 2653–2660, 1990.
Crampton, J., Humphries, S., Woods, D., and Williamson, R.: The isolation of clones cDNA sequences which are differetially expressed in human lymphocytes and fibroblasts. Nucleic Acids Res 8: 6007–6017, 1980.
Derman, E., Krauter, K., Walling, L., Weinberger, C., Ray, M., and Darnell, J., Jr.: Transcriptional control in the production of liver-specific mRNAs. Cell 23: 731–739, 1981.
Devereux, J., Haeberli, P., and Smithies, O.: A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res 12: 387–395, 1984.
Dworkin, M. and Dawid, I.: Construction of a cloned library of expressed embryonic gene sequences from Xenopus laevis. Dev Biol 76: 435–448, 1980.
Grunstein, M. and Hogness, D.: Colony hybridization: a method for the isolation of clones DNAs that contain a specific gene. Proc Natl Acad Sci USA 72: 3961–3965, 1975.
Hastie, N. and Bishop, J.: The expression of three abundance classes of messenger RNA in mouse tissues. Cell 9, 761–774, 1976.
Hochgeschwender, J., Sutcliffe, G., and Brennan, M.: Construction and screening of a genomic library specific for mouse chromosome 16. Proc Natl Acad Sci USA 86: 8482–8486, 1989.
Hoheisel, J.D., Lennon, G.G., Zehetner, G., and Lehrach, H.: Use of high coverage reference libraries of Drosophila melanogaster for relational data analysis. A step towards mapping and sequencing of the genome. J Mol Biol 220: 903–914, 1991a.
Hoheisel, J.D., Drmanac, R., Larin, Z., Lennon, G.G., Monaco, A., Zehetner, G., and Lehrach, H.: Use of high coverage libraries for an integrated analysis of genomic DNA. Adv Mol Genet 4: 125–132, 1991b.
Höög, C.: Isolation of a large number of novel mammalian genes by a differential cDNA library screening strategy. Nucleic Acids Res 19: 6123–6127, 1991.
Jacobs, H. and Birnie, G.D.: Post-transcriptional regulation of messenger abundance in rat liver and hepatoma. Nucleic Acids Res 8, 3087–3120, 1980.
Lehrach, H., Drmanac, R., Hoheisel, J., Larin, Z., Lennon, G., Monaco, A., Nizetic, D., Zehetner, G., and Poustka, A.: Hybridization fingerprinting in genome mapping and sequencing. In K. Davies and S.M. Tilghman (eds.); Genome Analysis, Vol. I: Genetic and Physical Mapping, pp. 39–81, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., 1990.
Lennon, G.G. and Lehrach, H.: Hybridization analyses of arrayed cDNA libraries. Trends Genet 7: 314–317, 1991.
Litt, M. and White, R.: A highly polymorphic locus in human DNA revealed by cosmid-derived probes. Proc Natl Acad Sci USA 82: 6206–6210, 1985.
Nizetic, D., Zehetner, G., Monaco, A., Gellen, L., Young B. and Lehrach, H.: Construction, arraying and high-density screening of large insert libraries of human chromosomes X and 21: their potential use as reference libraries. Proc Natl Acad Sci USA 88: 3233–3237, 1991.
Sambrook, J., Fritsch, E.F. and Maniatis, T.: Molecular Cloning, A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, 1989.
Sargent, T. and Dawid, I.: Differential gene expression in the gastrula of Xenopus laevis. Science 222: 135–139, 1983.
Sealy, P., Whittaker, P., and Southern, E.: Removal of repeated sequences from hybridisation probes. Nucleic Acids Res 13: 1905–1922, 1985.
Shiosaka, T. and Saunders, G.: Differential expression of selected genes in human leukemia leukocytes. Proc Natl Acad Sci USA 79: 4668–4671, 1982.
Shiosaka, T., Tanaka, Y., and Kobayashi, Y.: Preferentially expressed genes in stomach adenocarcinoma cells. Br J Cancer 56: 539–544, 1987.
Sutcliffe, J.G.: mRNA in the mammalian central nervous system. Annu Rev Neurosci 11: 157–198, 1988.
Sutcliffe, J.G., Milner, R.J., Bloom, F.J., and Lerner, R.: Common 82-nucleotide sequence unique to brain RNA. Proc Natl Acad Sci USA 79: 4942–4946, 1982.
Wang, D., Villasante, L.S., and Cowan, N.: The mammalian β-tubulin repertoire: hematopoietic expression of a novel heterologous β-tubulin isotype. J Cell Biol 103: 1903–1910, 1986.
Wiborg, O., Pedersen, M., Wind, A., Berglund, L., Marcker, K., and Vuust, J.: The human ubiquitin multigene family: some genes contain multiple directly repeated ubiquitin coding sequences. EMBO J 4: 755–759, 1985.
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Gress, T.M., Hoheisel, J.D., Lennon, G.G. et al. Hybridization fingerprinting of high-density cDNA-library arrays with cDNA pools derived from whole tissues. Mammalian Genome 3, 609–619 (1992). https://doi.org/10.1007/BF00352477
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DOI: https://doi.org/10.1007/BF00352477