Skip to main content
PMC home page
Cytotechnology logoLink to Cytotechnology
. 2006 Jul 25;50(1-3):121–140. doi: 10.1007/s10616-006-9004-9

Toward genomic cell culture engineering

Katie F Wlaschin 1, Gargi Seth 1, Wei-Shou Hu 1,✉
PMCID: PMC3476003  PMID: 19003075

Abstract

Genomic and proteomic based global gene expression profiling has altered the landscape of biological research in the past few years. Its potential impact on cell culture bioprocessing has only begun to emanate, partly due to the lack of genomic sequence information for the most widely used industrial cells, Chinese hamster ovary (CHO) cells. Transcriptome and proteome profiling work for species lacking extensive genomic resources must rely on information for other related species or on data obtained from expressed sequence tag (EST) sequencing projects, for which burgeoning efforts have only recently begun. This article discusses the aspects of EST sequencing in those industrially important, genomic resources-poor cell lines, articulates some of the unique features in employing microarray in the study of cultured cells, and highlights the infrastructural needs in establishing a platform for genomics based cell culture research. Recent experience has revealed that generally, most changes in culture conditions only elicit a moderate level of alteration in gene expression. Nevertheless, by broadening the conventional scope of microarray analysis to consider estimated levels of transcript abundance, much physiological insight can be gained. Examples of the application of microarray in cell culture are discussed, and the utility of pattern identification and process diagnosis are highlighted. As genomic resources continue to expand, the power of genomic tools in cell culture processing research will be amply evident. The key to harnessing the immense benefit of these genomic resources resides in the development of physiological understanding from their application.

Keywords: cDNA library, Data analysis, Mammalian cell culture, Microarray, Proteome, Transcriptome

Acknowledgements

The support from Pfizer, Inc. and Bayer Healthcare for the cell culture research work in W-S. Hu’s laboratory are gratefully acknowledged. KFW was supported by the NIH Biotechnology Training Grant (GM08347). The bioinformatic support was provided by The Minnesota Supercomputing Institute.

References

  1. Aebersold R, Mann M. Mass spectrometry-based proteomics. Nature. 2003;422:198–207. doi: 10.1038/nature01511. [DOI] [PubMed] [Google Scholar]
  2. Alete DE, Racher AJ, Birch JR, Stansfield SH, James DC, Smales CM. Proteomic analysis of enriched microsomal fractions from GS-NS0 murine myeloma cells with varying secreted recombinant monoclonal antibody productivities. Proteomics. 2005;5:4689–4704. doi: 10.1002/pmic.200500019. [DOI] [PubMed] [Google Scholar]
  3. Chakraborty A, Regnier FE. Global internal standard technology for comparative proteomics. J Chromatogr A. 2002;949:173–184. doi: 10.1016/S0021-9673(02)00047-X. [DOI] [PubMed] [Google Scholar]
  4. Chee M, Yang R, Hubbell E, Berno A, Huang XC, Stern D, Winkler J, Lockhart DJ, Morris MS, Fodor SP. Accessing genetic information with high-density DNA arrays. Science. 1996;274:610–614. doi: 10.1126/science.274.5287.610. [DOI] [PubMed] [Google Scholar]
  5. Chen Z, Southwick K, Thulin CD. Initial analysis of the phosphoproteome of Chinese hamster ovary cells using electrophoresis. J Biomol Tech: JBT. 2004;15:249–256. [PMC free article] [PubMed] [Google Scholar]
  6. Dahlquist KD, Salomonis N, Vranizan K, Lawlor SC, Conklin BR. GenMAPP, a new tool for viewing and analyzing microarray data on biological pathways. Nat Genet. 2002;1:19–20. doi: 10.1038/ng0502-19. [DOI] [PubMed] [Google Scholar]
  7. DeRisi JL, Iyer VR, Brown PO. Exploring the metabolic and genetic control of gene expression on a genomic scale. Science. 1997;278:681–685. doi: 10.1126/science.278.5338.680. [DOI] [PubMed] [Google Scholar]
  8. Durbin R, Eddy S, Krogh A, Mitchison G. Biological sequence analysis: probabilistic models of proteins and nucleic acids. Cambridge UK: Cambridge University Press; 1998. [Google Scholar]
  9. Gadgil C, Rink A, Beattie CW, Hu WS. A mathematical model for suppression subtractive hybridization. Comp Funct Genomics. 2002;3:405–422. doi: 10.1002/cfg.206. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Gadgil M, Lian W, Gadgil C, Kapur V, Hu W-S (2005) An analysis of the use of genomic DNA as a universal reference in two channel DNA microarrays. BMC Genomics 6 [DOI] [PMC free article] [PubMed]
  11. Gladney CD, Bertani GR, Johnson RK, Pomp D. Evaluation of gene expression in pigs selected for enhanced reproduction using differential display PCR and human microarrays: I. Ovarian follicles. J Anim Sci. 2004;82:17–31. doi: 10.2527/2004.82117x. [DOI] [PubMed] [Google Scholar]
  12. Gu J, Gu X. Induced gene expression in human brain after the split from chimpanzee. Trends Genet. 2003;19:63–65. doi: 10.1016/S0168-9525(02)00040-9. [DOI] [PubMed] [Google Scholar]
  13. Guo QM. DNA microarray and cancer. Curr Opin Oncol. 2003;15:36–43. doi: 10.1097/00001622-200301000-00005. [DOI] [PubMed] [Google Scholar]
  14. Gygi S, Rist B, Gerber S, Turecek F, Gelb M, Aebersold R. Quantitative analysis of complex protein mixtures using isotope-coded affinity tags. Nat Biotechnol. 1999;17:994–999. doi: 10.1038/13690. [DOI] [PubMed] [Google Scholar]
  15. Gygi SP, Corthals GL, Zhang Y, Rochon Y, Aebersold R. Evaluation of two-dimensional gel electrophoresis-based proteome analysis technology. Proc Natl Acad Sci USA. 2000;97:9390–9395. doi: 10.1073/pnas.160270797. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Harrison PM, Kumar A, Lang N, Snyder M, Gerstein M. A question of size: the eukaryotic proteome and the problems in defining it. Nucleic Acids Res. 2002;30:1083–1090. doi: 10.1093/nar/30.5.1083. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hayduk EJ, Choe LH, Lee KH. A two-dimensional electrophoresis map of Chinese hamster ovary cell proteins based on fluorescence staining. Electrophoresis. 2004;25:2545–2556. doi: 10.1002/elps.200406010. [DOI] [PubMed] [Google Scholar]
  18. Hayduk EJ, Lee KH. Cytochalasin D can improve heterologous protein productivity in adherent Chinese hamster ovary cells. Biotechnol Bioeng. 2005;90:354–364. doi: 10.1002/bit.20438. [DOI] [PubMed] [Google Scholar]
  19. Hunt DF, Henderson RA, Shabanowitz J, Sakaguchi K, Michel H, Sevilir N, Cox AL, Appella E, Engelhard VH. Characterization of peptides bound to the class I MHC molecule HLA-A2.1 by mass spectrometry. Science. 1992;255:1261–1263. doi: 10.1126/science.1546328. [DOI] [PubMed] [Google Scholar]
  20. International Human Genome Sequencing Consortium Initial sequencing and analysis of the human genome. Nature. 2001;409:860–921. doi: 10.1038/35057062. [DOI] [PubMed] [Google Scholar]
  21. Kaufmann H, Mazur X, Fussenegger M, Bailey JE. Influence of low temperature on productivity, proteome and protein phosphorylation of CHO cells. Biotechnol Bioeng. 1999;63:573–582. doi: 10.1002/(SICI)1097-0290(19990605)63:5<573::AID-BIT7>3.0.CO;2-Y. [DOI] [PubMed] [Google Scholar]
  22. Kim H, Zhao B, Snesrud E, Haas B, Town C, Quckenbush J. Use of RNA and genomic DNA references for inferred comparisons in DNA microarray analysis. Biotechniques. 2002;33:924–930. doi: 10.2144/02334mt06. [DOI] [PubMed] [Google Scholar]
  23. Korke R, DeLeon Gatti M, Lau AL, Lim JW, Seow TK, Chung MC, Hu WS. Large scale gene expression profiling of metabolic shift of mammalian cells in culture. J Biotechnol. 2004;107:1–17. doi: 10.1016/j.jbiotec.2003.09.007. [DOI] [PubMed] [Google Scholar]
  24. Lee MS, Kim KW, Kim YH, Lee GM. Proteome analysis of antibody-expressing CHO cells in response to hyperosmotic pressure. Biotechnol Prog. 2003;19:1734–1741. doi: 10.1021/bp034093a. [DOI] [PubMed] [Google Scholar]
  25. Lindblad-Toh K, Wade CM, Mikkelsen TS, Karlsson EK, Jaffe DB, Kamal M, Clamp M, Chang JL, Kulbokas EJ, Zody MC, et al (2005) Genome sequence, comparative analysis and haplotype structure of the domestic dog. 438:803–819 [DOI] [PubMed]
  26. Lockhart DJ, Dong H, Byrne MC, Follettie MT, Gallo MV, Chee MS, Mittmann M, Wang C, Kobayashi M, Horton H, Brown EL. Expression monitoring by hybridization to high-density oligonucleotide arrays. Nat Biotechnol. 1996;14:1675–1680. doi: 10.1038/nbt1296-1675. [DOI] [PubMed] [Google Scholar]
  27. Moody DE, Zou Z, McIntyre L. Cross-species hybridisation of pig RNA to human nylon microarrays. BMC Genomics. 2002;3:27. doi: 10.1186/1471-2164-3-27. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. MouseGenome Sequencing Consortium Initial sequencing and comparative analysis of the mouse genome. Nature. 2002;420:520–562. doi: 10.1038/nature01262. [DOI] [PubMed] [Google Scholar]
  29. Ong SE, Blagoev B, Kratchmarova I, Kristensen DB, Steen H, Pandey A, Mann M. Stable isotope labeling by amino acids in cell culture, SILAC, as a simple and accurate approach to expression proteomics. Mol Cell Proteomics. 2002;1:376–386. doi: 10.1074/mcp.M200025-MCP200. [DOI] [PubMed] [Google Scholar]
  30. Pattyn F, Speleman F, De Paepe A, Vandesompele J. RTPrimerDB: the real-time PCR primer and probe database. Nucleic Acids Res. 2003;31:122–123. doi: 10.1093/nar/gkg011. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Rabilloud T. Two-dimensional gel electrophoresis in proteomics: old, old fashioned, but it still climbs up the mountains. Proteomics. 2002;2:3–10. doi: 10.1002/1615-9861(200201)2:1<3::AID-PROT3>3.0.CO;2-R. [DOI] [PubMed] [Google Scholar]
  32. RatGenomeSequencing Project Consortium Genome sequence of the Brown Norway rat yields insights into mammalian evolution. Nature. 2004;428:493–521. doi: 10.1038/nature02426. [DOI] [PubMed] [Google Scholar]
  33. Rose K, Simona MG, Offord RE, Prior CP, Otto B, Thatcher DR. A new mass-spectrometric C-terminal sequencing technique finds a similarity between gamma-interferon and alpha 2-interferon and identifies a proteolytically clipped gamma-interferon that retains full antiviral activity. Biochem J. 1983;215:273–277. doi: 10.1042/bj2150273. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Ross P, Huang Y, Marchese J, Williamson B, Parker K, Hattan S, Khainovski N, Pillai S, Dey S, Daniels S, et al. Multiplexed protein quantitation in Saccharomyces cerevisiae using amine-reactive isobaric tagging reagents. Mol Cellular Proteomics. 2004;3:1154–1169. doi: 10.1074/mcp.M400129-MCP200. [DOI] [PubMed] [Google Scholar]
  35. Schneider LV, Hall MP. Stable isotope methods for high-precision proteomics. Drug Discovery Today. 2005;10:353–363. doi: 10.1016/S1359-6446(05)03381-7. [DOI] [PubMed] [Google Scholar]
  36. Seow TK, Korke R, Liang R, Ong S-E, Ou K, Wong K, Hu WS, Chung M. Proteomic investigation of metabolic shift in mammalian cell culture. Biotechnol Progr. 2001;17:1137–1144. doi: 10.1021/bp010101g. [DOI] [PubMed] [Google Scholar]
  37. Seth G, Philp RJ, Denoya CD, McGrath K, Stutzman-Engwall KJ, Yap MG, Hu W-S. Large scale gene expression analysis of cholesterol dependence in NS0 cells. Biotech Bioeng. 2005;90:552–567. doi: 10.1002/bit.20429. [DOI] [PubMed] [Google Scholar]
  38. Seth G, Ozturk M, Hu W-S (2006) Reverting cholesterol dependence of NS0 cells by altering epigenetic gene silencing. Biotech Bioeng 93:820–827 [DOI] [PubMed]
  39. Shen D, Sharfstein ST. Genome-wide analysis of the transcriptional response of murine hybridomas to osmotic shock. Biotechnol Bioeng. 2006;93:132–145. doi: 10.1002/bit.20691. [DOI] [PubMed] [Google Scholar]
  40. Smales CM, Dinnis DM, Stansfield SH, Alete D, Sage EA, Birch JR, Racher AJ, Marshall CT, James DC. Comparative proteomic analysis of GS-NS0 murine myeloma cell lines with varying recombinant monoclonal antibody production rate. Biotechnol Bioeng. 2004;88:474–488. doi: 10.1002/bit.20272. [DOI] [PubMed] [Google Scholar]
  41. Talaat A, Howard S, Hale W, Lyons R, Garner H, Johnston S (2002) Genomic DNA standards for gene expression profiling in Mycobacterium tuberculosis. Nucleic Acids Res 30 [DOI] [PMC free article] [PubMed]
  42. Unlu M, Morgan ME, Minden JS. Difference gel electrophoresis: a single gel method for detecting changes in protein extracts. Electrophoresis. 1997;18:2071–2077. doi: 10.1002/elps.1150181133. [DOI] [PubMed] [Google Scholar]
  43. Van Dyk DD, Misztal DR, Wilkins MR, Mackintosh JA, Poljak A, Varnai JC, Teber E, Walsh BJ, Gray PP. Identification of cellular changes associated with increased production of human growth hormone in a recombinant Chinese hamster ovary cell line. Proteomics. 2003;3:147–156. doi: 10.1002/pmic.200390023. [DOI] [PubMed] [Google Scholar]
  44. Williams B, Gwirtz R, Wold B (2004) Genomic DNA as a cohybridization standard for mammalian microarray measurements. Nucleic Acids Res 32 [DOI] [PMC free article] [PubMed]
  45. Wlaschin K, Nissom PM, Leon Gatti M, Fern PFO, Arleen S, Tan KS, Rink A, Cham B, Wong K, Yap M, Hu W-S. EST sequencing for gene discovery in Chinese hamster ovary cells. Biotech Bioeng. 2005;91:592–606. doi: 10.1002/bit.20511. [DOI] [PubMed] [Google Scholar]
  46. Wong VVT, Nissom PM, Sim S-L, Yeo JHM, Chuah S-H, Yap MGS. Zinc as an insulin replacement in hybridoma cultures. Biotechnol Bioeng. 2006;93:553–563. doi: 10.1002/bit.20746. [DOI] [PubMed] [Google Scholar]

Articles from Cytotechnology are provided here courtesy of Springer Science+Business Media B.V.

RESOURCES