Summary
Specific amplification of human sequences of up to several kb length has recently been accomplished in man-hamster and man-mouse somatic hybrid cell DNA by IRS-PCR (interspersed repetitive sequence — polymerase chain reaction). This approach is based on oligonucleotide primers that anneal specifically to human Alu- or L1-sequences and allows the amplification of any human sequences located between adequately spaced, inverted Alu- or L1-blocks. Here, we demonstrate that probe pools generated from two somatic hybrid cell lines by Alu- and L1-PCR can be used for chromosome painting in normal human lymphocyte metaphase spreads by chromosomal in situ suppression (CISS-) hybridization. The painted chromosomes and chromosome subregions directly represent the content of normal and deleted human chromosomes in the two somatic hybrid cell lines. The combination of IRS-PCR and CISS-hybridization will facilitate and improve the cytogenetic analysis of somatic hybrid cell panels, in particular, in cases where structurally aberrant human chromosomes or human chromosome segments involved in interspecies translocations cannot be unequivocally identified by classical banding techniques. Moreover, this new approach will help to generate probe pools for the specific delineation of human chromosome subregions for use in cytogenetic diagnostics and research without the necessity of cloning.
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References
Albertson DG, Fishpool R, Sherrington P, Nacheva E, Milstein C (1988) Sensitive and high resolution in situ hybridization to human chromosomes using biotin labelled probes: assignment of human thymocyte CD1 genes to chromosome 1. EMBO J 7:2801–2805
Bhatt B, Burns J, Flannery D, McGee O'D (1988) Direct visualization of single copy genes on banded metaphase chromosomes by non-isotopic in situ hybridization. Nucleic Acids Res 16:3951–3961
Boyle AL, Lichter P, Ward DC (1990) Rapid analysis of mousehamster hybrid cell lines by in situ hybridization. Genomics 7:127–130
Chen TL, Manuelidis L (1989) SINEs and LINEs cluster in distinct DNA fragments of Giemsa band size. Chromosoma 98:309–316
Cremer T, Lichter P, Borden J, Ward DC (1988) Detection of chromosome aberrations in metaphase and interphase tumor cells by in situ hybridization using chromosome-specific library probes. Hum Genet 80:235–246
Durnam DM, Gelinas RE, Myerson D (1985) Detection of species specific chromosomes in somatic cell hybrids. Somat Cell Mol Genet 11:571–577
Garson JA, Berghe JA van den, Kemshead JT (1987) Non isotopic in situ hybridization technique detects small (1kb) unique sequences in routinely G-banded human chromosomes: fine mapping of N-myc and β-NGF genes. Nucleic Acids Res 15:4761–4770
Kessler C, Höltke HJ, Seibl R, Burg J, Mühlegger K (1990) Nonisotopic labeling and detection of nucleic acids. I. A novel DNA labeling and detection system based on digoxigenin: antidigoxigenin ELISA principle (digoxigenin system). Biol Chem Hoppe-Seyler (in press)
Kievits T, Devilee P, Wiegant J, Wapenaar MC, Cornelisse CJ, Ommen GJB van, Pearson PL (1990) Direct nonradioactive in situ hybridization of somatic cell hybrid DNA to human lymphocyte chromosomes. Cytometry 11:105–109
Korenberg JR, Rykowski MC (1988) Human genome organization: Alu, Lines, and the molecular structure of metaphase chromosome bands. Cell 53:391–400
Landegent JE, Jansen in de Wal N, Dirks RW, Baas F, Ploeg M van der (1987) Use of whole cosmid clones genomic sequences for chromosomal localization by non-radioactive in situ hybridization. Hum Genet 77:366–370
Lawrence JB, Villnave CA, Singer RH (1988) Sensitive, high resolution chromatin and chromosome mapping in situ: presence and orientation of two closely integrated copies of EBV in a lymphoma line. Cell 52:51–61
Ledbetter SA, Nelson DL, Warren ST, Ledbetter DH (1990) Rapid isolation of DNA probes within specific chromosome regions by interspersed repetitive sequence (IRS) PCR. Genomics 6:475–481
Lengauer C, Eckelt A, Weith A, Endlich N, Ponelies N, Lichter P, Greulich KO, Cremer T (1990) Painting of defined chromosomal regions by in situ suppression hybridization of libraries from laser-microdissected chromosomes. Cytogenet Cell Genet (in press)
Lichter P, Ward DC (1990) Is non-isotopic in situ hybridization finally coming of age? Nature 345:93–95
Lichter P, Cremer T, Borden J, Manuelidis L, Ward DC (1988a) Delineation of individual human chromosomes in metaphase and interphase cells by in situ suppression hybridization using recombinant DNA libraries. Hum Genet 80:224–234
Lichter P, Cremer T, Tang CC, Watkins PC, Manuelidis L, Ward DC (1988b) Rapid detection of chromosome 21 aberrations by in situ hybridization. Proc Natl Acad Sci USA 85:9664–9668
Lichter P, Tang CC, Call C, Hermanson G, Evans GA, Housman D, Ward DC (1990) High-resolution mapping of human chromosome 11 by in situ hybridization with cosmid clones. Science 247:64–69
Maniatis T, Fritsch EF, Sambrook J (eds) (1982) Molecular cloning: a laboratory manual. Cold Spring Harbor Laboratory. Cold Spring Harbor, NY
Manuelidis L (1985) Individual interphase chromosome domains revealed by in situ hybridization. Hum Genet 71:288–293
Nelson DL, Ledbetter SA, Corbo L, Victoria MF, Ramirez-Solis R, Webster TD, Ledbetter DH, Caskey CT (1989) Alu polymerase chain reaction: a method for rapid isolation of humanspecific sequences from complex DNA sources. Proc Natl Acad Sci USA 86:6686–6690
Pinkel D, Gray JW, Trask B, Engh G van den. Fuscoe J, Dekken H van (1986) Cytogenetic analysis by in situ hybridization with fluorescently labeled nucleic acid probes. Cold Spring Harb Symp Quant Biol 51:151–157
Pinkel D, Landegent J, Collins C, Fuscoe J, Segraves R, Lucas J, Gray JW (1988) Fluorescence in situ hybridization with human chromosome specific libraries: detection of trisomy 21 and translocations of chromosome 4. Proc Natl Acad Sci USA 85:9138–9142
Ried T, Mahler V, Vogt P, Blonden L, Ommen GJB van, Cremer T, Cremer M (1990) Direct carrier detection by in situ suppression hybridization with cosmid clones of the Duchenne/Becker muscular dystrophy locus. Hum Genet 85:581–586
Riethman HC, Moyzis RK, Meyne J, Burke DT, Olson MV (1989) Cloning human telomeric DNA fragments into Saccharomyces cerevisiae using a yeast-artificial-chromosome vector. Proc Natl Acad Sci USA 86:6240–6244
Schardin M, Cremer T, Hager HD, Lang M (1985) Specific staining of human chromosomes in Chinese hamster x man hybrid ell lines. Hum Genet 71:281–287
Seibl R, Höltke HJ, Rüger R, Meindl A, Zachau HG, Raßhofer R, Roggendorf M, Wolf H, Arnold N, Wienberg J, Kessler C (1990) Non-radioactive labeling and detection of nucleic acids. III. Applications of the digoxigenin system. Biol Chem Hoppe Seyler (in press)
Trask B, Pinkel D, Engh G van den (1989) The proximity of DNA sequences in interphase cell nuclei is correlated to genomic distance and permits ordering of cosmids spanning 250 kilobase pairs. Genomics 5:710–717
Wada M, Little RD, Abidi F, Porta G, Labella T, Cooper T, Della Valle G, D'Urso M, Schlessinger D (1990) Human Xq24–Xq28: approaches to mapping with yeast artificial chromosomes. Am J Hum Genet 46:95–106
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Lengauer, C., Riethman, H. & Cremer, T. Painting of human chromosomes with probes generated from hybrid cell lines by PCR with Alu and L1 primers. Hum Genet 86, 1–6 (1990). https://doi.org/10.1007/BF00205163
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DOI: https://doi.org/10.1007/BF00205163