Abstract
It is now well recognized that chromosomal translocation followed by overexpression of a chimeric gene product plays a critical role in tumorigenicity in various malignant tumors, especially those of leukemia, malignant lymphoma, and soft-tissue tumors. In these malignant tumors, specific chimeric gene products are directly related to tumorigenicity. Therefore, if chimeric gene products could be observed in situ, it would be advantageous not only for the correct diagnosis of each tumor but also to improve our understanding of the basis of tumorigenicity. Accordingly, it would seem that reverse transcriptase (RT) in situ polymerase chain reaction (PCR) is a powerful and useful approach for the study of chimeric gene products in situ. Here, we introduce the application of RT in situ PCR to detect a hybrid, SYT-SSX messenger RNA in synovial sarcoma. We expect that the principle of this protocol also may be applied to detect other chimeric gene products.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Nuovo, G. J. (1996) PCR In situ Hybridization Protocols and Applications (2nd ed). Philadelphia, New York, Lippincott-Raven.
Nuovo, G. J. (1997) PCR In situ Hybridization Protocols and Applications (3rd ed). Philadelphia, New York, Lippincott-Raven.
Long, A. A., Komminoth, P., Lee, E., and Wolfe, H. F. (1993) Comparison of indirect and direct in-situ polymerase chain reaction in cell preparations and tissue sections. Histochemistry 99, 151–162.
Martinez, A., Miller, M. J., Quinn, K., Unsworth, E. J., Ebina, M., and Cuttitta, F. (1993) 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.
Komminoth, P. and Long, A. A. (1993) In-situ polymerase chain reaction: an overview of methods. Applications and limitations of a new molecular technique. Virchow Arch. (B) 64, 67–73
Wickham, C. L., Boyce, M., Joyner, M. V., et al. (2000) Amplification of PCR products in excess of 600 base pairs using DNA extracted from decalcified, paraffin wax embedded bone marrow trephine biopsies. Mol. Pathol. 53, 19–23.
Chiu, K. P., Cohen, S. H., Morris, D. W., and Jordan, G. W. (1992) Intracellular amplification of proviral DNA in tissue sections using the polymerase chain reaction. J. Histochem. Cytochem. 40, 333–341.
Clark, J., Rocques, P. J., Crew, A. J., et al. (1994) Identification of novel genes, SYT and SSX, involved in the t(X;18)(p11.2;q11.2) translocation found in human synovial sarcoma. Nat. Genet. 7, 502–508.
Crew, A. J., Clark, J., Fisher, C., et al. (1995) Fusion of STT to two genes, SSX1 and SSX2, encoding proteins with homology to the Kruppel-associated box in human synovial sarcoma. EMBO J. 14, 2333–2340.
Nagai, M., Tanaka, S., Tsuda, M., et al. (2001) Analysis of transforming activity of human synovial sarcoma-associated chimeric protein SYT-SSX1 bound to chromatin remodeling factor hBRM/hSNF2 alpha. Proc. Natl. Acad. Sci. USA. 98, 3843–3848.
Kato, H., Tjernberg, A., Zhang, W., et al. (2002) SYT associates with human SNF/SWI complexes and the C-terminal region of its fusion partner SSX1 targets histones. J. Biol. Chem. 277, 5498–5505.
Sonobe, H., Manabe, Y., Furihata, M., et al. (1992) Establishment and characterization of a new human synovial sarcoma cell line, HS-SY-II. Lab. Invest. 67, 98–505.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2006 Humana Press Inc.
About this protocol
Cite this protocol
Takeuchi, T., Adachi, Y., Sonobe, H., Ohtsuki, Y. (2006). Application of Reverse Transcription In Situ PCR in Cancer Analysis. In: Pellestor, F. (eds) PRINS and In Situ PCR Protocols. Methods in Molecular Biology™, vol 334. Humana Press. https://doi.org/10.1385/1-59745-068-5:169
Download citation
DOI: https://doi.org/10.1385/1-59745-068-5:169
Publisher Name: Humana Press
Print ISBN: 978-1-58829-549-1
Online ISBN: 978-1-59745-068-3
eBook Packages: Springer Protocols