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In Situ Reverse Transcription PCR for Detection of mRNA in the CNS

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Cellular and Molecular Methods in Neuroscience Research

Abstract

The development of polymerase chain reaction (PCR) produced a technological breakthrough in nucleic acid detection by increasing molecular sensitivity capabilities. In situ PCR is a marriage of two established technologies: PCR and in situ hybridization based on the amplification within intact cells or tissue sections of specific DNA sequences, or mRNA species, to levels detectable by in situ hybridization and/or immunohistochemistry. Thus, PCR results can be correlated spatially with cell morphology.

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References

  1. Bagasra, O., S.P. Hauptman, H.W. Lischner, M. Sachs, and R.J. Pomerantz. 1992. Detection of human immunodeficiency virus type 1 provirus in mononuclear cells by in situ polymerase chain reaction. N. Engl. J. Med. 326:1385–1391.

    Article  PubMed  CAS  Google Scholar 

  2. Chow, L.H., S. Subramanian, G.J. Nuovo, F. Miller, and E.P. Nord. 1995. Endothelin receptor mRNA expression in renal medulla identified by in situ RT-PCR. Am. J. Physiol. 269:F449–F457.

    PubMed  CAS  Google Scholar 

  3. Embretson, J., M. Zupancic, J. Beneke, M. Till, S. Wolinsky, J.L. Ribas, A. Burke, and A.T. Haase. 1993. Analysis of human immunodeficiency virus-infected tissues by amplification and in situ hybridization reveals latent and permissive infections at single-cell resolution. Proc. Natl. Acad. Sci. USA 90:357–361.

    Article  PubMed  CAS  Google Scholar 

  4. Frodin, M. and S. Gammeltoft. 1999. Role and regulation of 90 kDa ribosomal S6 kinase (RSK) in signal transduction. Mol. Cell. Endocrinol. 151:65–77.

    Article  PubMed  CAS  Google Scholar 

  5. Gu, J. 1995. In Situ Polymerase Chain Reaction and Related Technology. Eaton Publishing, Natick, MA.

    Google Scholar 

  6. Haase, A.T., E.F. Retzel, and K.A. Staskus. 1990. Amplification and detection of lentiviral DNA inside cells. Proc. Natl. Acad. Sci. USA 87:4971–4975.

    Article  PubMed  CAS  Google Scholar 

  7. Herrington, C.S. and J.J. O’Leary. 1998. PCR In Situ Hybridization. Oxford University Press, New York.

    Google Scholar 

  8. Jacob, A., I.R. Hurley, L.O. Goodwin, G.W. Cooper, and S. Benoff. 2000. Molecular characterization of a voltage-gated potassium channel expressed in rat testis. Mol. Hum. Reprod. 6:303–313.

    Article  PubMed  CAS  Google Scholar 

  9. Jacquot, S., K. Merienne, E. Trivier, M. Zeniou, S. Pannetier, and A. Hanauer. 1999. Coffin-Lowry syndrome: current status. Am. J. Med. Genet. 85:214–215.

    Article  PubMed  CAS  Google Scholar 

  10. Jensen, C.J., M.B. Buch, T.O. Krag, B.A. Hemmings, S. Gammeltoft, and M. Frodin. 1999. 90-kDa ribosomal S6 kinase is phosphorylated and activated by 3-phosphoinositide-dependent protein kinase-1. J. Biol. Chem. 274:27168–27176.

    Article  PubMed  CAS  Google Scholar 

  11. Koffron, A.J., M. Hummel, B.K Patterson, S. Yan, D.B. Kaufman, J.P. Fryer, F.P. Stuart, and M.I. Abecassis. 1998. Cellular localization of latent murine cytomegalovirus. J. Virol. 72:95–103.

    PubMed  CAS  Google Scholar 

  12. Komminoth, P. and A.A. Long. 1993. In-situ polymerase chain reaction. An overview of methods, applications and limitations of a new molecular technique. Virchows Arch. B Cell Pathol. Ind. Mol. Pathol. 64:67–73.

    CAS  Google Scholar 

  13. Komminoth, P., A.A. Long, R. Ray, and H.J. Wolfe. 1992. In situ polymerase chain reaction detection of viral DNA, single-copy genes, and gene rearrangements in cell suspensions and cytospins. Diagn. Mol. Pathol. 1:85–97.

    Article  PubMed  CAS  Google Scholar 

  14. Kulaksiz, H., R. Arnold, B. Goke, E. Maronde, M. Meyer, F. Fahrenholz, W.G. Forssmann, and R. Eissele. 2000. Expression and cell-specific localization of the cholecystokinin B/gastrin receptor in the human stomach. Cell Tissue Res. 299:289–298.

    Article  PubMed  CAS  Google Scholar 

  15. Lewis, F. 1996. An Approach to In situ PCR. PE Biosystems, Foster City, CA.

    Google Scholar 

  16. Long, A.A. 1998. In-situ polymerase chain reaction: foundation of the technology and today’s options. Eur. J. Histochem. 42:101–109.

    PubMed  CAS  Google Scholar 

  17. Long, A.A. and P. Komminoth. 1997. In situ PCR. An overview. Methods Mol. Biol. 71:141–161.

    PubMed  CAS  Google Scholar 

  18. Long, A.A., P. Komminoth, E. Lee, and H.J. Wolfe. 1993. Comparison of indirect and direct in-situ polymerase chain reaction in cell preparations and tissue sections. Detection of viral DNA, gene rearrangements and chromosomal translocations. Histochemistry 99:151–162.

    Article  PubMed  CAS  Google Scholar 

  19. Martinez, A., M.J. Miller, K Quinn, E.J. Unsworth, M. Ebina, and F. Cuttitta. 1995. Non-radioactive localization of nucleic acids by direct in situ PCR and in situ RT-PCR in paraffin-embedded sections. J. Histochem. Cytochem. 43:739–747.

    PubMed  CAS  Google Scholar 

  20. Nakai, M., T. Kawamata, T. Taniguchi, K Maeda, and C. Tanaka. 1996. Expression of apolipoprotein E mRNA in rat microglia. Neurosci. Lett. 211:41–44.

    Article  PubMed  CAS  Google Scholar 

  21. Nuovo, G.J. 2000. In situ localization of PCR-amplified DNA and cDNA. Methods Mol. Biol. 123:217–238.

    PubMed  CAS  Google Scholar 

  22. Nuovo, G.J. 1994. PCR In Situ Hybridization. Raven Press, New York.

    Book  Google Scholar 

  23. Nuovo, G.J., F. Gallery, P. MacConnell, J. Becker, and W Bloch. 1991. An improved technique for the in situ detection of DNA after polymerase chain reaction amplification. Am. J. Pathol. 139:1239–1244.

    PubMed  CAS  Google Scholar 

  24. Ohtaka-Maruyama, C., F. Hanaoka, and A.B. Chepelinsky. 1998. A novel alternative spliced variant of the transcription factor AP2alpha is expressed in the murine ocular lens. Dev. Biol. 202:125–135.

    Article  PubMed  CAS  Google Scholar 

  25. O’Leary, J.J., R. Chetty, A.K. Graham, and J.O. McGee. 1996. In situ PCR: pathologist’s dream or nightmare? J. Pathol. 178:11–20.

    Article  PubMed  CAS  Google Scholar 

  26. Patterson, B.K., M. Till, P. Otto, C. Goolsby, M.R. Furtado, L.J. McBride, and S.M. Wolinsky. 1993. Detection of HIV-1 DNA and messenger RNA in individual cells by PCR-driven in situ hybridization and flow cytometry. Science 260:976–979.

    Article  PubMed  CAS  Google Scholar 

  27. Trivier, E., D. De Cesare, S. Jacquot, S. Pannetier, E. Zackai, I. Young, J.L. Mandel, P. Sassone-Corsi, and A. Hanauer. 1996. Mutations in the kinase Rsk-2 associated with Coffin-Lowry syndrome. Nature 384:567–570.

    Article  PubMed  CAS  Google Scholar 

  28. Wilcox, J.N. 1993. Fundamental principles of in situ hybridization. J. Histochem. Cytochem. 41:1725–1993.

    PubMed  CAS  Google Scholar 

  29. Wolfahrt, S., B. Heine, and W.G. Rossmanith. 1998. Detection of gonadotrophin releasing hormone and its receptor mRNA in human placental trophoblasts using in-situ reverse transcription-polymerase chain reaction. Mol. Hum. Reprod. 4:999–1006.

    Article  PubMed  CAS  Google Scholar 

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Author information

Authors and Affiliations

  1. Department of Neuropathology, Rigshospitalet Copenhagen University Hospital, Copenhagen

    Helle Broholm

  2. Department of Clinical Biochemistry, Glostrup Hospital, Copenhagen University, Glostrup, Denmark, EU

    Steen Gammeltoft

Authors
  1. Helle Broholm
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  2. Steen Gammeltoft
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Editor information

Editors and Affiliations

  1. Department of Veterinary Morphophysiology Rita Levi Montalcini Center Brain Repair, University of Torino, Gruglasco, I-10055, Italy

    Adalberto Merighi

  2. Department of Experimental and Biomedical Sciences CNR Center for the Study of Biomembranes, University of Padova, Padova, I-35121, Italy

    Giorgio Carmignoto

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© 2002 Springer-Verlag New York, Inc.

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Broholm, H., Gammeltoft, S. (2002). In Situ Reverse Transcription PCR for Detection of mRNA in the CNS. In: Merighi, A., Carmignoto, G. (eds) Cellular and Molecular Methods in Neuroscience Research. Springer, New York, NY. https://doi.org/10.1007/978-0-387-22460-2_9

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  • DOI: https://doi.org/10.1007/978-0-387-22460-2_9

  • Publisher Name: Springer, New York, NY

  • Print ISBN: 978-0-387-95386-1

  • Online ISBN: 978-0-387-22460-2

  • eBook Packages: Springer Book Archive

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