Use of whole cosmid cloned genomic sequences for chromosomal localization by non-radioactive in situ hybridization (original) (raw)

Summary

We report a general procedure which allows the application of whole cosmid cloned genomic sequences for non-radioactive in situ hybridization. The presence of highly repetitive sequences, like Alu and Kpn fragments, is eliminated through competition hybridization with Cot-1 DNA. The method has been tested and optimized with several randomly chosen cosmids of the human thyroglobulin (Tg) gene (8q24). At present, the procedure can be performed with three of the four tested individual cosmids. In cases where a single clone does not result in a specific signal, a larger fragment may be required, which can be accomplished by using two (partially overlapping) cosmids of the same region. The advantages and further potentialities of such a hybridization approach are discussed.

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References

• Albertson DG (1985) Mapping muscle protein genes by in sity hybridization using biotin-labeled probes. EMBO J 4:2493–2498
  Google Scholar
• Ambros PF, Matzke MA, Matzke AJM (1986) Detection of a 17 kb unique sequence (T-DNA) in plant chromosomes by in situ hybridization. Chromosoma 94:11–18
  Google Scholar
• Baas F, Bikker H, Geurts van Kessel A, Melsert R, Pearson PL, De Vijlder JJM, Van Ommen G-JB (1985) The human thyroglobulin gene: a polymorphic marker localized distal to c-myc on chromosome 8 band q24. Hum Genet 69:138–143
  Google Scholar
• Baas F, Van Ommen G-JB, Bikker H, Arnberg AC, De Vijlder JJM (1986) The human thyroglobulin gene is over 300 kb long and contains introns of up to 64 kb. Nucleic Acids Res 14:5171–5186
  Google Scholar
• Brison O, Ardeshir F, Stark GR (1982) General method for amplified DNA by differential screening with genomic probes. Mol Cell Biol 2:578–587
  Google Scholar
• Burns J, Chan VTW, Jonasson JA, Fleming KA, Taylor S, McGee JO'D (1985) Sensitive system for visualising biotinylated DNA probes hybridised in situ: rapid sex determination of infact cells. J Clin Pathol 38:1085–1092
  Google Scholar
• Cremer T, Landegent JE, Bruckner H, Scholl HP, Schardin M, Hager HD, Devilee P, Pearson PL, Van der Ploeg M (1986) Detection of chromosome aberrations in the human interphase nucleus by visualization of specific target DNAs with radioactive and non-radioactive in situ hybridization techniques: diagnosis of trisomy 18 with probe L1.84. Hum Genet 74:346–352
  Google Scholar
• Cremers F, Jansen in de Wal N, Wiegant J, Dirks RW, Weisbeek P, Van der Ploeg M, Landegent JE (1987) Non-radioactive in situ hybridization: a comparison of several immunocytochemical detection systems using reflection-contrast and electron microscopy. Histochemistry 86:600–615
  Google Scholar
• De Pagter-Holthuizen P, Van Schaik FMA, Verduijn GM, Van Ommen G-JB, Bouma BN, Jansen M, Sussenbach JS (1986) Organization of the human genes for insulin-like growth factors I and II. FEBS Lett 195:179–184
  Google Scholar
• Fisher JH, Gusella JF, Scoggin CH (1984) Molecular hybridization under conditions of high stringency permits cloned DNA segments containing reiterated DNA sequences to be assigned to specific chromosomal locations. Proc Natl Acad Sci USA 81:520–524
  Google Scholar
• Fuchs R, Daune M (1972) Physical studies on deoxyribonucleic acid after covalent binding of a carcinogen. Biochemistry 11:2659–2666
  Google Scholar
• Gerhard DS, Kawasaki ES, Bancroft FC, Szabo P (1981) Localization of a unique gene by direct hybridization in situ. Proc Natl Acad Sci USA 78:3755–3759
  Google Scholar
• Harper ME, Saunders GF (1981) Localiztion of single-copy DNA sequences of G-banded human chromosomes by in situ hybridization. Chromosoma 83:431–439
  Google Scholar
• Harper ME, Ullrich A, Saunders GF (1981) Localization of the human insulin gene to the distal end of the short arm of chromosome 11. Proc Natl Acad Sci USA 78:4458–4460
  Google Scholar
• Hopman AHN, Wiegant J, Van Duijn P (1986a) A new hybridocytochemical method based on mercurated nucleic acid probes and sulfhydryl-hapten ligands. I. Stability of the mercury-sulfhydryl bond and influence of the ligand structure on immunochemical detection of the hapten. Histochemistry 84:169–185
  Google Scholar
• Hopman AHN, Wiegant J, Tesser GI, Van Duijn P (1986b) A nonradioactive in situ hybridization method based on mercurated nucleic acid probes and sulfhydryl-hapten ligands. Nucleic Acids Res 14:6471–6488
  Google Scholar
• Landegent JE, Jansen in de Wal N, Baan RA, Hoeijmakers JHJ, Van der Ploeg M (1984) 2-Acetylaminofluorene-modified probes for the indirect hybridocytochemical detection of specific nucleic aicd sequences. Exp Cell Res 153:61–72
  Google Scholar
• Landegent JE, Jansen in de Wal N, Ploem JS, Van der Ploeg M (1985a) Sensitive detection of hybridocytochemical results by means of reflection-contrast microscopy. J Histochem Cytochem 33:1241–1246
  Google Scholar
• Landegent JE, Jansen in de Wal N, Van Ommen G-JB, Baas F, De Vijlder JJM, Van Duijn P, Van der Ploeg M (1985b) Chromosomal localization of a unique gene by non-autoradiographic in situ hybridization. Nature 317:175–177
  Google Scholar
• Landegent JE, Jansen in de Wal N, Fisser-Groen YM, Bakker E, Van der Ploeg M, Pearson PL (1986) Fine mapping of the Huntington disease linked D4S10 locus by non-radioactive in situ hybridization. Hum Genet 73:354–357
  Google Scholar
• Langer-Safer PR, Levine M, Ward DC (1982) Immunological method for mapping genes on Drosophila polytene chromosomes. Proc Acad Sci USA 79:4381–4385
  Google Scholar
• Ploem JS (1975) Reflection-contrast microscopy as a tool for the investigation of the attachment of the living cells to glass surface. In: Van Furth R (ed) Mononuclear phagocytes in immunity, infection and pathology. Blackwell, Oxford London, pp 405–421
  Google Scholar
• Schmid CW, Jelinek WR (1982) The Alu family of dispersed repetitive sequences. Science 216:1065–1070
  Google Scholar
• Sealey PG, Whittaker PA, Southern EM (1985) Removal of repeated sequences from hybridization probes. Nucleic Acids Res 13: 1905–1922
  Google Scholar
• Shroyer KR, Nakane PK (1983) Use of DNP-labeled cDNA for in situ hybridization. J Cell Biol 97: 377a
• Singer MF (1982) SINEs and LINEs: highly repeated short and long interspersed sequences in mammalian genomes. Cell 28:433–434
  Google Scholar
• Tchen P, Fuchs RPP, Sage E, Leng M (1984) Chemically modified nucleic acids as immunodetectable probes in hybridization experiments. Proc Natl Acad Sci USA 81:3466–3570
  Google Scholar
• Trask BJ, Van den Engh G, Landegent JE, Jansen in de Wal N, Van der Ploeg M (1985) Detection of DNA sequences in nuclei in suspension by in situ hybridization and dual beam flow cytometry. Science 230:1401–1403
  Google Scholar
• Ullu E (1982) The human Alu family of repeated DNA sequences. Trends Biochem Sci 7:216–219
  Google Scholar
• Van Ommen G-JB, Arnberg AC, Baas F, Brocas H, Sterk A, Tegelaers HH, Vassart G, De Vijlder JJM (1983) The human thyroglobulin gene contains two 15–17 kb introns near its 3′-end. Nucleic Acids Res 11:2273–2285
  Google Scholar
• Weiss EH, Golden L, Fahrner K, Mellor AL, Devlin JJ, Bullman H, Tiddens H, Bud H, Flavell RA (1984) Organization and evolution of class I gene family in the major histocompatibility complex of the C57BL/10 mouse. Nature 310:650–555
  Google Scholar

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Author notes

1. J. E. Landegent
   Present address: Biomedical Sciences Division, Lawrence Livermore National Laboratory, P.O. Box 5507, 94550, Livermore, CA, USA
2. R. W. Dirks
   Present address: Department of Biochemistry and Molecular Biology, Harvard University, 7 Divinity Avenue, 02138, Cambridge, MA, USA

Authors and Affiliations

1. Department of Cytochemistry and Cytometry, Sylvius Laboratories, University of Leiden, P.O. Box 9503, NL-2333 AL, Leiden, The Netherlands
   J. E. Landegent, N. Jansen in de Wal, R. W. Dirks & M. van der Ploeg
2. Division of Molecular Biology, The Netherlands Cancer Institute (Antonie van Leeuwenhoekhuis), Plesmanlaan 121, NL-1066 CX, Amsterdam, The Netherlands
   R. W. Dirks

Authors

1. J. E. Landegent
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2. N. Jansen in de Wal
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3. R. W. Dirks
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4. M. van der Ploeg
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Landegent, J.E., Jansen in de Wal, N., Dirks, R.W. et al. Use of whole cosmid cloned genomic sequences for chromosomal localization by non-radioactive in situ hybridization.Hum Genet 77, 366–370 (1987). https://doi.org/10.1007/BF00291428

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• Received: 28 April 1987
• Issue Date: December 1987
• DOI: https://doi.org/10.1007/BF00291428

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