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A recombination–based assay demonstrates that the fragile X sequence is transcribed widely during development

Nature Genetics volume 3, pages 44–48 (1993)Cite this article

Abstract

To identify transcribed sequences rapidly and efficiently, we have developed a recombination–based assay to screen bacteriophage λ libraries for sequences that share homology with a given probe. This strategy determines analytically whether a given probe is transcribed in a given tissue at a given time of development, and may also be used to isolate preparatively the transcribed sequence free of the screening probe. We illustrate this technology for the fragile X sequence, demonstrating that it is transcribed ubiquitously in an 11 week fetus, in a variety of 20 week human fetal tissues, including brain, spinal cord, eye, liver, kidney and skeletal muscle, and in adult jejunum

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References

  1. Watt, V.M., Ingles, C.J., Urdea, M.S. & Rutter, W.J. Homology requirements for recombination in Escherichia coli. Proc. natn. Acad. Sci. U.S.A. 82, 4768–4772 (1985).

    Article  CAS  Google Scholar 

  2. Shen, P. & Huang, H.V. Homologous recombination in Escherichia coli: dependence on substrate length and homology. Genetics 112, 441–457 (1986).

    CAS  PubMed  PubMed Central  Google Scholar 

  3. King, S.R. & Richardson, J.P. Role of homology and pathway specificity for recombination between plasmids and bacteriophage λ. Molec. gen. Genet. 204, 141–147, (1986).

    Article  CAS  Google Scholar 

  4. Shen, P. & Huang, H.V. Effect of base pair mismatches on recombination via the RecBCD pathway. Molec. gen. Genet. 218, 358–360 (1989).

    Article  CAS  Google Scholar 

  5. Verkerk, A.J. et al. Identification of a gene (FMR-1) containing a CGG repeat coincident with a breakpoint cluster region exhibiting length variation in fragile X syndrome. Cell 65, 905–914 (1991).

    Article  CAS  Google Scholar 

  6. Pieretti, M. et al. Absence of expression of the FMR-1 gene in fragile X syndrome. Cell 66, 817–822 (1991).

    Article  CAS  Google Scholar 

  7. Stewart, G.D. et al. Plasmids for recombination-based screening. Gene 106, 97–101 (1991).

    Article  CAS  Google Scholar 

  8. Hanzlik, A.J., Osemlak, M.M. & Kurnit, D.M. A small plasmid for recombination-based screening. Gene (in the press).

  9. Nisson, P.E., Rashtchian, A. & Watkins, P.C. Rapid and efficient cloning of Alu-PCR products using uracil DNA glycosylase. PCR Meth. Applic. 1, 120–123 (1991).

    Article  CAS  Google Scholar 

  10. Seed, B. Purification of genomic sequences from bacteriophage libraries by recombination and selection in vivo. Nucl. Acids Res. 11, 2427–2445 (1983).

    Article  CAS  Google Scholar 

  11. Kurnit, D.M. & Seed, B. Improved genetic selection for screening bacteriophage libraries by homologous recombination in vivo. Proc. natn. Acad. Sci. U.S.A. 87, 3166–3169 (1990).

    Article  CAS  Google Scholar 

  12. Georgopoulos, C. & Herskowitz, I. in The Bacteriophage Lambda, (ed. Hershey, A.D.) 553–563 (Cold Spring Harbor, New York, 1971).

    Google Scholar 

  13. Saiki, R.K. et al. Enzymatic amplification of β-globin genomic sequences and restriction site analysis for diagnosis of sickle cell anemia. Science 230, 1350–1354 (1985).

    Article  CAS  Google Scholar 

  14. Fu, Y.H. et al. Variation of the CGG repeat at the fragile X site results in genetic instability: resolution of the Sherman paradox. Cell 67, 1047–1058 (1991).

    Article  CAS  Google Scholar 

  15. Neve, R.L. et al. Human chromosome 21 -encoded cDNA clones. Gene 49, 361–369 (1986).

    Article  CAS  Google Scholar 

  16. Short, J.M., Fernandez, J.M., Sorge, J.A. & Huse, W.D. λZAP: a bacteriophage λ expression vector with in vivo excision properties. Nucl. Acids Res. 16, 7583–7600 (1985).

    Article  Google Scholar 

  17. Uberbacher, E.G. & Mural, R.J. Locating protein-coding regions in human DNA sequences by a multiple sensor-neural network approach. Proc. natn. Acad. Sci. U.S.A. 88, 11261–11265 (1991).

    Article  CAS  Google Scholar 

  18. Liu, P., Legerski, R. & Siciliano, M.J. Isolation of human transcribed sequences from human-rodent somatic cell hybrids. Science 246, 813–815 (1989).

    Article  CAS  Google Scholar 

  19. Duyk, G.M., Kim, S., Myers, R.M. & Cox, D.R. Exon trapping: a genetic screen to identify candidate transcribed sequences in cloned mammalian genomic DNA. Proc. natn. Acad. Sci. U.S.A. 87, 8995–8999 (1990).

    Article  CAS  Google Scholar 

  20. Buckler, A.J. et al. Exon amplificaton: a strategy to isolate mammalian genes based on RNA splicing. Proc. natn. Acad. Sci. U.S.A. 88, 4005–4009 (1991).

    Article  CAS  Google Scholar 

  21. Lovett, M., Kere, J. & Hinton, L.M. Direct selection: a method for the isolation of cDNAs encoded by large genomic regions. Proc. natn. Acad. Sci. U.S.A. 88, 9628–9632 (1991).

    Article  CAS  Google Scholar 

  22. Parimoo, S., Patanjali, S.R., Shukla, H., Chaplin, D.D. & Weissman, S.M. cDNA selection: efficient PCR approach for the selection of cDNAs encoded in large chromosomal DNA fragments. Proc. natn. Acad. Sci. U.S.A. 88, 9623–9627 (1991).

    Article  CAS  Google Scholar 

  23. Adams, M.D. et al. Complementary DNA sequencing: expressed sequence tags and human genome project. Science 252, 1651–1656 (1991).

    Article  CAS  Google Scholar 

  24. Guarente, L., Lauer, G., Roberts, T.M. & Ptashne, M. Improved methods for maximizing expression of a cloned gene: a bacterium that synthesizes rabbit β-globin. Cell 20, 543–553 (1980).

    Article  CAS  Google Scholar 

  25. Casadaban, M.J. & Cohen, S.N. Analysis of gene control signals by DNA fusion and cloning in Escherichia coli. J. molec. Biol. 138, 179–207 (1980).

    Article  CAS  Google Scholar 

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Authors and Affiliations

  1. University of Michigan Medical School, Howard Hughes Medical Institute, Ann Arbor Michigan, 48109-0650, USA

    Andrzej J. Hanzlik, Malgorzata M. Osemlak-Hanzlik, Michael A. Hauser & David M. Kurnit

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  1. Andrzej J. Hanzlik
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  2. Malgorzata M. Osemlak-Hanzlik
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  3. Michael A. Hauser
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  4. David M. Kurnit
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Hanzlik, A., Osemlak-Hanzlik, M., Hauser, M. et al. A recombination–based assay demonstrates that the fragile X sequence is transcribed widely during development. Nat Genet 3, 44–48 (1993). https://doi.org/10.1038/ng0193-44

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