Summary
Since its invention in 1992, differential display (DD) has become the most commonly used technique for identifying differentially expressed genes because of its many advantages over competing technologies such as DNA microarray, serial analysis of gene expression (SAGE), and subtractive hybridization. A large number of these publications have been in the field of cancer, specifically on p53 target genes. Despite the great impact of the method on biomedical research, there had been a lack of automation of DD technology to increase its throughput and accuracy for systematic gene expression analysis. Many previous DD work has taken a “shotgun” approach of identifying one gene at a time, with a limited number of polymerase chain reactions (PCRs) set up manually, giving DD a low-tech and low-throughput image. We have optimized the DD process with a platform that incorporates fluorescent digital readout, automated liquid handling, and large-format gels capable of running entire 96-well plates. The resulting streamlined fluorescent DD (FDD) technology offers an unprecedented accuracy, sensitivity, and throughput in comprehensive and quantitative analysis of gene expression. These major improvements will allow researchers to find differentially expressed genes of interest, both known and novel, quickly and easily.
Key words
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Sager, R. (1997) Expression genetics in cancer: shifting the focus from DNA to RNA. Proc. Natl. Acad. Sci. USA 94,952–955.
Vogelstein, B., Lane, D., and Levine, A.J. (2000) Surfing the p53 network. Nature 408, 307–310.
Liang, P., and Pardee, A.B. (1992) Differential display of eukaryotic messenger RNA by means of the polymerase chain reaction. Science 257, 967–971.
Liang, P. (2002) A decade of differential display. Biotechniques 33, 338–346.
Liang, P., and Pardee, A.B. (2003) Analysing differential gene expression in cancer. Nat. Rev. Cancer 3, 869–876.
Liang, P., Meade, J., and Pardee, A.B. (2007) A protocol for differential display of mRNA expression using either fluorescent or radioactive labeling. Nat. Protoc.2, 457–470.
Schena, M., Shalon, D., Davis, R.W., and Brown, P.O. (1995) Quantitative monitoring of gene expression patterns with a complementary DNA microarray. Science 270, 467–470.
Chee, M., Yang, R., Hubbell, E., Berno, A., Huang, X.C., Stern, D., et al. (1996) Accessing genetic information with high-density DNA arrays. Science274, 610–614.
Velculescu V.E., Zhang L., Vogelstein B., and Kinzler K.W. (1995) Serial analysis of gene expression. Science 270, 484–487.
Zimmermann, C.R., Orr, W.C., Leclerc, R.F., Barnard, E.C., and Timberlake, W.E. (1980) Molecular cloning and selection of genes regulated in Aspergillus development. Cell21, 709–715.
McCarthy, S.A., Samuels, M.L., Pritchard, C.A., Abraham, J.A., and McMahon, M. (1995) Rapid induction of heparin-binding epidermal growth factor/diphtheria toxin receptor expression by Raf and Ras oncogenes. Genes Devel. 9, 1953–1964.
Zhang, R., Tan, Z., and Liang, P. (2000) Identification of a novel ligand-receptor pair constitutively activated by Ras oncogenes.J. Biol. Chem. 275, 24436–24443.
You, M., Ku, P.T., Hrdlickova, R., and Bose, H.R., Jr. (1997) ch-IAP1, a member of the inhibitor-of-apoptosis protein family, is a mediator of the antiapoptotic activity of the v-Rel oncoprotein. Mol. Cell. Biol. 17, 7328–7341.
Park, B.-W., O’Rourke, D.M., Wang, Q., Davis, J.G., Post, A., Qian, X., et al. (1999) Induction of the Tat-binding protein 1 gene accompanies the disabling of oncogenic erbB receptor tyrosine kinases. Proc. Natl. Acad. Sci. USA 96, 6434–6438.
Wang, M., Tan, Z., Zhang, R., Kotenko, S.V., and Liang, P. (2002) Interleukin-24 (Mob-5/Mda-7) signals through two heterodimeric receptors, IL-22R1/IL-20R2 and IL-20R1/IL-20R2. J. Biol. Chem. 277, 7341–7347.
El-Deiry, W.S. (1998) Regulation of p53 downstream genes. Semin. Cancer Biol. 8, 345–357.
Wu, X., Bayle, J.H., Olson, D., and Levine, A.J. (1993) The p53-mdm-2 autoregulatory feedback loop. Genes Dev. 7, 1126–1132.
El-Deiry W.S., Tokino, T., Velculescu, V.E., Levy, D.B., Parsons, R., Trent, J.M., et al. (1993) WAF1, a potential mediator of p53 tumor suppression. Cell 75, 817–825.
Miyashita, T., and Reed, J.C. (1995) Tumor suppressor p53 is a direct transcriptional activator of the human bax gene. Cell 80, 293–299
Okamoto, K., and Beach, D. (1994) Cyclin G is a transcriptional target of the p53 tumor suppressor protein. EMBO J. 13, 4816–4822.
Buckbinder, L, Talbott, R., Velasco-Miguel, S., Takenaka, I., Faha, B., Seizinger, B.R., et al. (1995) Induction of the growth inhibitor IGF-binding protein 3 by p53. Nature 377, 646–649.
Polyak, K., Xia, Y., Zweier, J.L, Kinzler, K.W., and Vogelstein, B. (1997) A model for p53-induced apoptosis. Nature 389, 300–305.
Wu, G.S., Burns, T.F., McDonald, E.R., Jiang, W., Meng, R., Krantz, I.D., et al. (1997) KILLER/DR5 is a DNA damage-inducible p53-regulated death receptor gene. Nat. Genet. 17, 141–143.
Gu, Z., Flemington, C., Chittenden, T., and Zambetti, G.P. (2000) ei24, a p53 response gene involved in growth suppression and apoptosis. Mol. Cell. Biol. 20, 233–241.
Israeli, D., Tessler, E., Haupt, Y., Elkeles, A., Wilder, S., Amson, R., et al. (1997) A novel p53-inducible gene, PAG608, encodes a nuclear zinc finger protein whose overexpression promotes apoptosis. EMBO J. 16, 4384–4392.
Lo, P.K., Chen, J.-Y., Lo, W.-C., Chen, B.-F., Hsin, J.-P., Tang, P.-P, et al. (1999) Identification of a novel mouse p53 target gene DDA3. Oncogene 18, 7765–7774.
Takei, Y., Ishikawa, S., Tokino, T., Muto, T., and Nakamura, Y. (1998) Isolation of a novel TP53 target gene from a colon cancer cell line carrying a highly regulated wild-type TP53 expression system. Genes Chromosomes Cancer 23, 1–9.
Ng, C.C., Koyama, K., Okamura, S., Kondoh, H., Takei, Y., and Nakamura, Y. (1999) Isolation and characterization of a novel TP53-inducible gene, TP53TG3. Genes Chromosomes Cancer 26, 329–335.
Tanaka, H., Arakawa, H., Yamaguchi, T., Shiraishi, K., Fukuda, S., Matsui, K., et al. (2000) A ribonucleotide reductase gene involved in a p53-dependent cell-cycle checkpoint for DNA damage. Nature 404, 42–49.
Attardi, L., Reczek, E.E., Cosmas, C., Demicco, E.G., McCurrach, M.E., Lowe, S.W., et al. (2000) PERP, an apoptosis-associated target of p53, is a novel member of the PMP-22/gas3 family. Genes Dev. 14, 704–718.
Saller, E., Tom, E., Brunori, M., Otter, M., Estreicher, A., Mack, D.H., et al. (1999) Increased apoptosis induction by 121F mutant p53. EMBO J. 18, 4424–4437.
Oda, E., Ohki, R., Murasawa, H., Nemoto, J., Shibue, T., Yamashita, T., et al. (2000) Noxa, a BH3-only member of the Bcl-2 family and candidate mediator of p53-induced apoptosis. Science 288, 1053–1058.
Lin, Y., Ma, W., and Benchimol, S. (2000) Pidd, a new death-domain-containing protein is induced by p53 and promotes apoptosis. Nat. Genet. 26, 124–127.
Oda, E., Arakawa, H., Tanaka, T., Matsuda, K., Tanikawa, C., Mori, T., et al. (2000) p53AIP1, a potential mediator of p53-dependent apoptosis, and its regulation by Ser-46-phosphorylated p53. Cell 102, 849–862.
Okamura, S., Arakawa, H., Tanaka, T., Nakanishi, H., Ng, C.C., Taya, Y., et al. (2001) p53DINP1, a p53-inducible gene, regulates p53-dependent apoptosis. Mol. Cell 8, 85–94.
Yu, J., Zhang, L, Hwang, P.M., Kinzler, K.W., and Vogelstein, B. (2001) PUMA induces the rapid apoptosis of colorectal cancer cells. Mol. Cell 7, 673–682.
Nakano, K., and Vousden, K.H. (2001) PUMA, a novel proapoptotic gene, is induced by p53. Mol. Cell 7, 683–694.
Leng, R.P., Lin, Y., Ma, W., Wu, H., Lemmers, B., Chung, S., et al. (2003) Pirh2, a p53-induced ubiquitin-protein ligase, promotes p53 degradation. Cell 112, 779–791.
Yin, Y., Liu, Y.-X., Jin, Y.J., Hall, E.J., and Barrett, J.C. (2003) PAC1 phosphatase is a transcription target of p53 in signalling apoptosis and growth suppression. Nature 422, 527–531.
Owen-Schaub, L.B., Zhang, W., Cusack, J.C., Angelo, L.S., Santee, S.M., Fujiwara, T., et al. (1995) Wild-type human p53 and a temperature-sensitive mutant induce Fas/APO-1 expression. Mol. Cell. Biol. 15, 3032–3040.
Kannan, K., Kaminski, N., Rechavi, G., Jakob-Hirsch, J., Amariglio, N., and Givol, D. (2001) DNA microarray analysis of genes involved in p53 mediated apoptosis: activation of Apaf-1. Oncogene 20, 3449–3455.
Stambolic, V., MacPherson, D., Sas, D., Lin, Y., Snow, B., Jang, Y., et al. (2001) Regulation of PTEN transcription by p53. Mol. Cell 8, 317–325.
Sax, J.K., Fei, P., Murphy, M.E., Bernhard, E., Korsmeyer, S.J., and El-Deiry, W.S. (2002) BID regulation by p53 contributes to chemosensitivity. Nat. Cell Biol. 411, 842–849.
Cho, Y.-J., Meade, J.D., Walden, J.C., Chen, X., Guo, Z., and Liang, P. (2001) Multicolor fluorescent differential display. Biotechniques 30, 562–572.
Meade, J.D., Cho, Y.-J., Fisher, J.S., Walden, J.C., Guo, Z., and Liang, P. (2005) Automation of fluorescent differential display with digital readout. In Differential Display Methods and Protocols, 2nd edition. Vol. 317 (Liang, P., Meade, J.D., & Pardee, A.B., eds.) Humana Press, Totowa, NJ, pp. 23–57.
Bauer, D., Muller, H., Reich, J., Riedel, H., Ahrenkiel, V., Warthoe, P., et al. (1993) Identification of differentially expressed mRNA species by an improved display technique (DDRT-PCR). Nucleic Acids Res. 21, 4272–4280.
Liang, P., Bauer, D., Averboukh, L., Warthoe, P., Rohrwild, M., Muller, H., et al. (1995) Analysis of altered gene expression by differential display. Methods Enzymol. 254, 304–321.
Liang, P., Zhu, W., Zhang, X., Guo, Z., O’Conell, R.P., Averboukh, L., et al. (1994) Differential Display using one-base anchored oligo-dT primers. Nucleic Acids Res. 22, 5763–5764.
Liang, P., Averboukh, L., and Pardee, A.B. (1994) Method of differential display. In Methods in Molecular Genetics, (Adolph, K.W., ed.) Academic, San Diego, CA, pp. 3–16.
Yang, S., and Liang, P. (2004) Global analysis of gene expression by differential display - a mathematical model. Mol. Biotechnol. 27, 197–208.
Liang, P., Averboukh, L., and Pardee, A.B. (1993) Distribution and cloning of eukaryotic mRNAs by means of differential display: Refinements and optimization. Nucleic Acids Res. 21, 3269–3275.
Hsu, D.K., Donohue, P.J., Alberts, G.F., and Winkles, J.A. (1993) Fibroblast growth factor-1 induces phosphofructokinase, fatty acid synthase and Ca (2+)-ATPase mRNA expression in NIH 3T3 cells. Biochem. Biophys. Res. Commun. 197, 1483–1491.
Sokolov, B.P., and Prockop, D.J. (1994) A rapid and simple PCR-based method for isolation of cDNAs from differentially expressed genes. Nucleic Acids Res. 22, 4009–4015.
Irie, T., Oshida, T., Hasegawa, H., Matsuoka, Y., Li, T., Oya, Y., et al. (2000) Automated DNA fragment collection by capillary array gel electrophoresis in search of differentially expressed genes. Electrophoresis 21, 367–374.
Ausubel, F., Brent, R., Kingston, R. E., Moore, D. D., Seidman, J. G., Smith, J. A., et al. (eds.) (1995) Short Protocols in Molecular Biology (3rd edition). Wiley, New York, NY. Section 4.9.1.–4.9.8.
Zhang, H., Zhang, R., and Liang, P. (1996) Differential screening of gene expression difference enriched by differential display. Nucleic Acids Res. 24, 2454–2455.
Ramdas, L., Coombes, K.R., Baggerly, K., Abruzzo, L., Highsmith, W.E., Krogmann, T., et al. (2001) Sources of nonlinearity in cDNA microarray expression measurements. Genome Biol. 2, RESEARCH0047.
Richmond, C.S., Glasner, J.D., Mau, R., Jin, H., and Blattner, F.R. (1999) Genome-wide expression profiling in Escherichia coli K-12. Nucleic Acids Res. 27, 3821–3835.
Gibbs, W.W. (2001) Shrinking to enormity: DNA microarrays are reshaping basic biology – but scientist fear that they may soon drown in data. Sci. Am. 284, 33–34.
Liang, P. (2000) Gene discovery using differential display. Gen. Eng. News 20, 37.
Liang, S., Rossby, S.P., Liang, P., Shelton, R.C., Manier, D.H., Chakrabarti, A., et al. (2005) Detection of an mRNA polymorphism by differential display. In Differential Display Methods and Protocols, 2nd edition. Vol. 317 (Liang, P., Meade, J.D., & Pardee, A.B., eds.) Humana Press, Totowa, NJ, pp 279–285.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2009 Humana Press, a part of Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Meade, J.D., Cho, Yj., Shester, B.R., Walden, J.C., Guo, Z., Liang, P. (2009). Automated Fluorescent Differential Display for Cancer Gene Profiling. In: Grützmann, R., Pilarsky, C. (eds) Cancer Gene Profiling. Methods in Molecular Biology, vol 576. Humana Press. https://doi.org/10.1007/978-1-59745-545-9_7
Download citation
DOI: https://doi.org/10.1007/978-1-59745-545-9_7
Published:
Publisher Name: Humana Press
Print ISBN: 978-1-934115-76-3
Online ISBN: 978-1-59745-545-9
eBook Packages: Springer Protocols