Skip to main content
SpringerLink
Log in

Methods in mammalian cell line engineering: from random mutagenesis to sequence-specific approaches

  • Mini-Review
  • Published:
Applied Microbiology and Biotechnology Aims and scope Submit manuscript

Abstract

Due to the increasing demand for recombinant proteins, the interest in mammalian cell culture, especially of Chinese hamster ovary cells, grows rapidly. This is accompanied by the desire to improve cell lines in order to achieve higher titers and a better product quality. Until recently, most cell line development procedures were based on random integration and gene amplification, but several methods for targeted genetic modification of cells have been developed. Some of those are homologous recombination, RNA interference and zinc-finger nucleases. Especially the latter two have evolved considerably and will soon become a standard for cell line engineering in research and industrial application. This review presents an overview of established as well as new and promising techniques for targeted genetic modification of mammalian cells.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3

Similar content being viewed by others

References

  • Al-Rubeai M, Singh RP (1998) Apoptosis in cell culture. Curr Opin Biotechnol 9(2):152–156

    Article  CAS  Google Scholar 

  • Aravin AA, Hannon GJ, Brennecke J (2007) The Piwi-piRNA pathway provides an adaptive defense in the transposon arms race. Science 318(5851):761–764

    Article  CAS  Google Scholar 

  • Beerli RR, Barbas CF 3rd (2002) Engineering polydactyl zinc-finger transcription factors. Nat Biotechnol 20(2):135–141

    Article  CAS  Google Scholar 

  • Bernstein E, Caudy AA, Hammond SM, Hannon GJ (2001) Role for a bidentate ribonuclease in the initiation step of RNA interference. Nature 409(6818):363–366

    Article  CAS  Google Scholar 

  • Bi JX, Shuttleworth J, Al-Rubeai M (2004) Uncoupling of cell growth and proliferation results in enhancement of productivity in p21CIP1-arrested CHO cells. Biotechnol Bioeng 85(7):741–749

    Article  CAS  Google Scholar 

  • Bibikova M, Beumer K, Trautman JK, Carroll D (2003) Enhancing gene targeting with designed zinc finger nucleases. Science 300(5620):764

    Article  CAS  Google Scholar 

  • Bibikova M, Golic M, Golic KG, Carroll D (2002) Targeted chromosomal cleavage and mutagenesis in Drosophila using zinc-finger nucleases. Genetics 161(3):1169–1175

    CAS  Google Scholar 

  • Birzele F, Schaub J, Rust W, Clemens C, Baum P, Kaufmann H, Weith A, Schulz TW, Hildebrandt T (2010) Into the unknown: expression profiling without genome sequence information in CHO by next generation sequencing. Nucleic Acids Res 38(12):3999–4010

    Article  CAS  Google Scholar 

  • Brummelkamp TR, Bernards R, Agami R (2002) A system for stable expression of short interfering RNAs in mammalian cells. Science 296(5567):550–553

    Article  CAS  Google Scholar 

  • Carthew RW, Sontheimer EJ (2009) Origins and mechanisms of miRNAs and siRNAs. Cell 136(4):642–655

    Article  CAS  Google Scholar 

  • Castanotto D, Sakurai K, Lingeman R, Li H, Shively L, Aagaard L, Soifer H, Gatignol A, Riggs A, Rossi JJ (2007) Combinatorial delivery of small interfering RNAs reduces RNAi efficacy by selective incorporation into RISC. Nucleic Acids Res 35(15):5154–5164

    Article  CAS  Google Scholar 

  • Cathomen T, Joung JK (2008) Zinc-finger nucleases: the next generation emerges. Mol Ther 16(7):1200–1207

    Article  CAS  Google Scholar 

  • Choudhuri S (2009) Lesser known relatives of miRNA. Biochem Biophys Res Commun 388(2):177–180

    Article  CAS  Google Scholar 

  • Cost GJ, Freyvert Y, Vafiadis A, Santiago Y, Miller JC, Rebar E, Collingwood TN, Snowden A, Gregory PD (2010) BAK and BAX deletion using zinc-finger nucleases yields apoptosis-resistant CHO cells. Biotechnol Bioeng 105(2):330–340

    Article  CAS  Google Scholar 

  • Durai S, Mani M, Kandavelou K, Wu J, Porteus MH, Chandrasegaran S (2005) Zinc finger nucleases: custom-designed molecular scissors for genome engineering of plant and mammalian cells. Nucleic Acids Res 33(18):5978–5990

    Article  CAS  Google Scholar 

  • Dykxhoorn DM, Novina CD, Sharp PA (2003) Killing the messenger: short RNAs that silence gene expression. Nat Rev Mol Cell Biol 4(6):457–467

    Article  CAS  Google Scholar 

  • Elbashir SM, Harborth J, Lendeckel W, Yalcin A, Weber K, Tuschl T (2001) Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature 411(6836):494–498

    Article  CAS  Google Scholar 

  • Epinat JC, Arnould S, Chames P, Rochaix P, Desfontaines D, Puzin C, Patin A, Zanghellini A, Pâques F, Lacroix E (2003) A novel engineered meganuclease induces homologous recombination in yeast and mammalian cells. Nucleic Acids Res 31(11):2952–2962

    Article  CAS  Google Scholar 

  • Figueroa B Jr, Ailor E, Osborne D, Hardwick JM, Reff M, Betenbaugh MJ (2007) Enhanced cell culture performance using inducible anti-apoptotic genes E1B-19K and Aven in the production of a monoclonal antibody with Chinese hamster ovary cells. Biotechnol Bioeng 97(4):877–892

    Article  CAS  Google Scholar 

  • Figueroa B Jr, Chen S, Oyler GA, Hardwick JM, Betenbaugh MJ (2004) Aven and Bcl-xL enhance protection against apoptosis for mammalian cells exposed to various culture conditions. Biotechnol Bioeng 85(6):589–600

    Article  CAS  Google Scholar 

  • Fire A, Xu S, Montgomery MK, Kostas SA, Driver SE, Mello CC (1998) Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans. Nature 391(6669):806–811

    Article  CAS  Google Scholar 

  • Foley JE, Yeh JR, Maeder ML, Reyon D, Sander JD, Peterson RT, Joung JK (2009) Rapid mutation of endogenous zebrafish genes using zinc finger nucleases made by Oligomerized Pool ENgineering (OPEN). PLoS ONE 4(2):e4348

    Article  CAS  Google Scholar 

  • Fukuda MN, Sasaki H, Lopez L, Fukuda M (1989) Survival of recombinant erythropoietin in the circulation: the role of carbohydrates. Blood 73(1):84–89

    CAS  Google Scholar 

  • Fussenegger M, Schlatter S, Dätwyler D, Mazur X, Bailey JE (1998) Controlled proliferation by multigene metabolic engineering enhances the productivity of Chinese hamster ovary cells. Nat Biotechnol 16(5):468–472

    Article  CAS  Google Scholar 

  • Golden DE, Gerbasi VR, Sontheimer EJ (2008) An inside job for siRNAs. Mol Cell 31(3):309–312

    Article  CAS  Google Scholar 

  • Goswami J, Sinskey AJ, Steller H, Stephanopoulos GN, Wang DI (1999) Apoptosis in batch cultures of Chinese hamster ovary cells. Biotechnol Bioeng 62(6):632–640

    Article  CAS  Google Scholar 

  • Grabher C, Wittbrodt J (2007) Meganuclease and transposon mediated transgenesis in medaka. Genome Biol 8 Suppl. 1:S10

    Article  Google Scholar 

  • Grimm D, Streetz KL, Jopling CL, Storm TA, Pandey K, Davis CR, Marion P, Salazar F, Kay MA (2006) Fatality in mice due to oversaturation of cellular microRNA/short hairpin RNA pathways. Nature 441(7092):537–541

    Article  CAS  Google Scholar 

  • Grizot S, Epinat JC, Thomas S, Duclert A, Rolland S, Pâques F, Duchateau P (2010) Generation of redesigned homing endonucleases comprising DNA-binding domains derived from two different scaffolds. Nucleic Acids Res 38(6):2006–2018

    Article  CAS  Google Scholar 

  • Grizot S, Smith J, Daboussi F, Prieto J, Redondo P, Merino N, Villate M, Thomas S, Lemaire L, Montoya G, Blanco FJ, Pâques F, Duchateau P (2009) Efficient targeting of a SCID gene by an engineered single-chain homing endonuclease. Nucleic Acids Res 37(16):5405–5419

    Article  CAS  Google Scholar 

  • Hou Y, Gao F, Wang Q, Zhao J, Flagg T, Zhang Y, Deng X (2007) Bcl2 impedes DNA mismatch repair by directly regulating the hMSH2-hMSH6 heterodimeric complex. J Biol Chem 282(12):9279–9287

    Article  CAS  Google Scholar 

  • Ifandi V, Al-Rubeai M (2005) Regulation of cell proliferation and apoptosis in CHO-K1 cells by the coexpression of c-Myc and Bcl-2. Biotechnol Prog 21(3):671–677

    Article  CAS  Google Scholar 

  • Imai-Nishiya H, Mori K, Inoue M, Wakitani M, Iida S, Shitara K, Satoh M (2007) Double knockdown of alpha1, 6-fucosyltransferase (FUT8) and GDP-mannose 4, 6-dehydratase (GMD) in antibody-producing cells: a new strategy for generating fully non-fucosylated therapeutic antibodies with enhanced ADCC. BMC Biotechnol 7:84

    Article  CAS  Google Scholar 

  • Irani N, Beccaria AJ, Wagner R (2002) Expression of recombinant cytoplasmic yeast pyruvate carboxylase for the improvement of the production of human erythropoietin by recombinant BHK-21 cells. J Biotechnol 93(3):269–282

    Article  CAS  Google Scholar 

  • Izsvák Z, Chuah MK, Vandendriessche T, Ivics Z (2006) Efficient stable gene transfer into human cells by the Sleeping Beauty transposon vectors. Methods 49(3):287–297

    Article  CAS  Google Scholar 

  • Jackson AL, Linsley PS (2010) Recognizing and avoiding siRNA off-target effects for target identification and therapeutic application. Nat Rev Drug Discov 9(1):57–67

    Article  CAS  Google Scholar 

  • Jayapal KP, Wlaschin KF, Hu WS, Yap MGS (2007) Recombinant protein therapeutics from CHO cells—20 years and counting. Chem Eng Prog 103(10):40–47

    CAS  Google Scholar 

  • Kanda Y, Imai-Nishiya H, Kuni-Kamochi R, Mori K, Inoue M, Kitajima-Miyama K, Okazaki A, Iida S, Shitara K, Satoh M (2007) Establishment of a GDP-mannose 4, 6-dehydratase (GMD) knockout host cell line: a new strategy for generating completely non-fucosylated recombinant therapeutics. J Biotechnol 130(3):300–310

    Article  CAS  Google Scholar 

  • Kandavelou K, Ramalingam S, London V, Mani M, Wu J, Alexeev V, Civin CI, Chandrasegaran S (2009) Targeted manipulation of mammalian genomes using designed zinc finger nucleases. Biochem Biophys Res Commun 388(1):56–61

    Article  CAS  Google Scholar 

  • Kantardjieff A, Nissom PM, Chuah SH, Yusufi F, Jacob NM, Mulukutla BC, Yap M, Hu WS (2009) Developing genomic platforms for Chinese hamster ovary cells. Biotechnol Adv 6:1028–1035

    Article  CAS  Google Scholar 

  • Kao FT, Puck TT (1968) Genetics of somatic mammalian cells, VII. Induction and isolation of nutritional mutants in Chinese hamster cells. Proc Natl Acad Sci USA 60(4):1275–1281

    Article  CAS  Google Scholar 

  • Kim DH, Rossi JJ (2007) Strategies for silencing human disease using RNA interference. Nat Rev Genet 8(3):173–184

    Article  CAS  Google Scholar 

  • Kim SH, Lee GM (2007) Down-regulation of lactate dehydrogenase-A by siRNAs for reduced lactic acid formation of Chinese hamster ovary cells producing thrombopoietin. Appl Microbiol Biotechnol 74(1):152–159

    Article  CAS  Google Scholar 

  • Lee EU, Roth J, Paulson JC (1989) Alteration of terminal glycosylation sequences on N-linked oligosaccharides of Chinese hamster ovary cells by expression of beta-galactoside alpha 2, 6-sialyltransferase. J Biol Chem 264(23):13848–13855

    CAS  Google Scholar 

  • Lee Y, Ahn C, Han J, Choi H, Kim J, Yim J, Lee J, Provost P, Rådmark O, Kim S, Kim VN (2003) The nuclear RNase III Drosha initiates microRNA processing. Nature 425(6956):415–419

    Article  CAS  Google Scholar 

  • Lee YY, Wong KT, Tan J, Toh PC, Mao Y, Brusic V, Yap MG (2009) Overexpression of heat shock proteins (HSPs) in CHO cells for extended culture viability and improved recombinant protein production. J Biotechnol 143(1):34–43

    Article  CAS  Google Scholar 

  • Lim SF, Chuan KH, Liu S, Loh SO, Chung BY, Ong CC, Song Z (2006) RNAi suppression of Bax and Bak enhances viability in fed-batch cultures of CHO cells. Metab Eng 8(6):509–522

    Article  CAS  Google Scholar 

  • Liu PQ, Chan EM, Cost GJ, Zhang L, Wang J, Miller JC, Guschin DY, Reik A, Holmes MC, Mott JE, Collingwood TN, Gregory PD (2010) Generation of a triple-gene knockout mammalian cell line using engineered zinc-finger nucleases. Biotechnol Bioeng 106(1):97–105

    CAS  Google Scholar 

  • Maeder ML, Thibodeau-Beganny S, Osiak A, Wright DA, Anthony RM, Eichtinger M, Jiang T, Foley JE, Winfrey RJ, Townsend JA, Unger-Wallace E, Sander JD, Müller-Lerch F, Fu F, Pearlberg J, Göbel C, Dassie JP, Pruett-Miller SM, Porteus MH, Sgroi DC, Iafrate AJ, Dobbs D, McCray PB Jr, Cathomen T, Voytas DF, Joung JK (2008) Rapid “open-source” engineering of customized zinc-finger nucleases for highly efficient gene modification. Mol Cell 31(2):294–301

    Article  CAS  Google Scholar 

  • Mani M, Smith J, Kandavelou K, Berg JM, Chandrasegaran S (2005) Binding of two zinc finger nuclease monomers to two specific sites is required for effective double-strand DNA cleavage. Biochem Biophys Res Commun 334(4):1191–1197

    Article  CAS  Google Scholar 

  • Mastrangelo AJ, Hardwick JM, Zou S, Betenbaugh MJ (2000) Part II. Overexpression of bcl-2 family members enhances survival of mammalian cells in response to various culture insults. Biotechnol Bioeng 67(5):555–564

    Article  CAS  Google Scholar 

  • Matasci M, Hacker DL, Baldi L, Wurm FM (2008) Recombinant therapeutic protein production in cultivated mammalian cells: current status and future prospects. Drug Discovery Today: Technol 5(2–3):e37–e42

    Article  Google Scholar 

  • May T, Hauser H, Wirth D (2006) Current status of transcriptional regulation systems. Cytotechnology 50(1–3):109–119

    Article  CAS  Google Scholar 

  • Miller JC, Holmes MC, Wang J, Guschin DY, Lee YL, Rupniewski I, Beausejour CM, Waite AJ, Wang NS, Kim KA, Gregory PD, Pabo CO, Rebar EJ (2007) An improved zinc-finger nuclease architecture for highly specific genome editing. Nat Biotechnol 25(7):778–785

    Article  CAS  Google Scholar 

  • Minch SL, Kallio PT, Bailey JE (1995) Tissue plasminogen activator coexpressed in Chinese hamster ovary cells with alpha(2, 6)-sialyltransferase contains NeuAc alpha(2, 6)Gal beta(1, 4)Glc-N-AcR linkages. Biotechnol Prog 11(3):348–351

    Article  CAS  Google Scholar 

  • Miyoshi H, Blömer U, Takahashi M, Gage FH, Verma IM (1998) Development of a self-inactivating lentivirus vector. J Virol 72(10):8150–8157

    CAS  Google Scholar 

  • Mori K, Kuni-Kamochi R, Yamane-Ohnuki N, Wakitani M, Yamano K, Imai H, Kanda Y, Niwa R, Iida S, Uchida K, Shitara K, Satoh M (2004) Engineering Chinese hamster ovary cells to maximize effector function of produced antibodies using FUT8 siRNA. Biotechnol Bioeng 88(7):901–908

    Article  CAS  Google Scholar 

  • Müller D, Katinger H, Grillari J (2008) MicroRNAs as targets for engineering of CHO cell factories. Trends Biotechnol 26(7):359–365

    Article  CAS  Google Scholar 

  • Nehlsen K, Broll S, Bode J (2006) Replicating minicircles: generation of nonviral episomes for the efficient modification of dividing cells. Gene Ther Mol Biol 10:233–244

    Google Scholar 

  • Ngantung FA, Miller PG, Brushett FR, Tang GL, Wang DI (2006) RNA interference of sialidase improves glycoprotein sialic acid content consistency. Biotechnol Bioeng 95(1):106–119

    Article  CAS  Google Scholar 

  • O’Callaghan PM, James DC (2008) Systems biotechnology of mammalian cell factories. Brief Funct Genomic Proteomic 7(2):95–110

    Article  CAS  Google Scholar 

  • Ohya T, Hayashi T, Kiyama E, Nishii H, Miki H, Kobayashi K, Honda K, Omasa T, Ohtake H (2008) Improved production of recombinant human antithrombin III in Chinese hamster ovary cells by ATF4 overexpression. Biotechnol Bioeng 100(2):317–324

    Article  CAS  Google Scholar 

  • Omasa T, Takami T, Ohya T, Kiyama E, Hayashi T, Nishii H, Miki H, Kobayashi K, Honda K, Ohtake H (2008) Overexpression of GADD34 enhances production of recombinant human antithrombin III in Chinese hamster ovary cells. J Biosci Bioeng 106(6):568–573

    Article  CAS  Google Scholar 

  • Oumard A, Qiao J, Jostock T, Li J, Bode J (2006) Recommended method for chromosome exploitation: RMCE-based cassette-exchange systems in animal cell biotechnology. Cytotechnology 50(1–3):93–108

    Article  CAS  Google Scholar 

  • Pâques F, Duchateau P (2007) Meganucleases and DNA double-strand break-induced recombination: perspectives for gene therapy. Curr Gene Ther 7(1):49–66

    Article  Google Scholar 

  • Rémy S, Tesson L, Ménoret S, Usal C, Scharenberg AM, Anegon I (2010) Zinc-finger nucleases: a powerful tool for genetic engineering of animals. Transgenic Res 19(3):363–371

    Article  CAS  Google Scholar 

  • Santiago Y, Chan E, Liu PQ, Orlando S, Zhang L, Urnov FD, Holmes MC, Guschin D, Waite A, Miller JC, Rebar EJ, Gregory PD, Klug A, Collingwood TN (2008) Targeted gene knockout in mammalian cells by using engineered zinc-finger nucleases. Proc Natl Acad Sci USA 105(15):5809–5814

    Article  Google Scholar 

  • Sauerwald TM, Betenbaugh MJ, Oyler GA (2002) Inhibiting apoptosis in mammalian cell culture using the caspase inhibitor XIAP and deletion mutants. Biotechnol Bioeng 77(6):704–716

    Article  CAS  Google Scholar 

  • Schlenke P, Grabenhorst E, Nimtz M, Conradt HS (1999) Construction and characterization of stably transfected BHK-21 cells with human-type sialylation characteristic. Cytotechnology 30(1–3):17–25

    Article  CAS  Google Scholar 

  • Schwarz DS, Hutvágner G, Du T, Xu Z, Aronin N, Zamore PD (2003) Asymmetry in the assembly of the RNAi enzyme complex. Cell 115(2):199–208

    Article  CAS  Google Scholar 

  • Seth G, Hossler P, Yee JC, Hu WS (2006) Engineering cells for cell culture bioprocessing—physiological fundamentals. Adv Biochem Eng Biotechnol 101:119–164

    CAS  Google Scholar 

  • Singh RP, Finka G, Emery AN, Al-Rubeai M (1997) Apoptosis and its control in cell culture systems. Cytotechnology 23:87–93

    Article  Google Scholar 

  • Siomi H, Siomi MC (2009) On the road to reading the RNA-interference code. Nature 457(7228):396–404

    Article  CAS  Google Scholar 

  • Spearman M, Rodriguez J, Huzel N, Butler M (2005) Production and glycosylation of recombinant beta-interferon in suspension and cytopore microcarrier cultures of CHO cells. Biotechnol Prog 21(1):31–39

    Article  CAS  Google Scholar 

  • Sung YH, Lee JS, Park SH, Koo J, Lee GM (2007) Influence of co-down-regulation of caspase-3 and caspase-7 by siRNAs on sodium butyrate-induced apoptotic cell death of Chinese hamster ovary cells producing thrombopoietin. Metab Eng 9(5–6):452–464

    Article  CAS  Google Scholar 

  • Tjio JH, Puck TT (1958) Genetics of somatic mammalian cells. II. Chromosomal constitution of cells in tissue culture. J Exp Med 108(2):259–268

    Article  CAS  Google Scholar 

  • Tomari Y, Zamore PD (2005) Perspective: machines for RNAi. Genes Dev 19(5):517–529

    Article  CAS  Google Scholar 

  • Urlaub G, Chasin LA (1980) Isolation of Chinese hamster cell mutants deficient in dihydrofolate reductase activity. Proc Natl Acad Sci USA 77(7):4216–4220

    Article  CAS  Google Scholar 

  • Urlaub G, Käs E, Carothers AM, Chasin LA (1983) Deletion of the diploid dihydrofolate reductase locus from cultured mammalian cells. Cell 33(2):405–412

    Article  CAS  Google Scholar 

  • Urnov FD, Miller JC, Lee YL, Beausejour CM, Rock JM, Augustus S, Jamieson AC, Porteus MH, Gregory PD, Holmes MC (2005) Highly efficient endogenous human gene correction using designed zinc-finger nucleases. Nature 435(7042):646–651

    Article  CAS  Google Scholar 

  • Vasquez KM, Marburger K, Intody Z, Wilson JH (2001) Manipulating the mammalian genome by homologous recombination. Proc Natl Acad Sci USA 98(15):8403–8410

    Article  CAS  Google Scholar 

  • Wang Q, Gao F, May WS, Zhang Y, Flagg T, Deng X (2008) Bcl2 negatively regulates DNA double-strand-break repair through a nonhomologous end-joining pathway. Mol Cell 29(4):488–498

    Article  CAS  Google Scholar 

  • Wlaschin KF, Nissom PM, Gatti Mde L, Ong PF, Arleen S, Tan KS, Rink A, Cham B, Wong K, Yap M, Hu WS (2005) EST sequencing for gene discovery in Chinese hamster ovary cells. Biotechnol Bioeng 91(5):592–606

    Article  CAS  Google Scholar 

  • Wolfe SA, Nekludova L, Pabo CO (2000) DNA recognition by Cys2His2 zinc finger proteins. Annu Rev Biophys Biomol Struct 29:183–212

    Article  CAS  Google Scholar 

  • Wong DC, Wong KT, Lee YY, Morin PN, Heng CK, Yap MG (2006a) Transcriptional profiling of apoptotic pathways in batch and fed-batch CHO cell cultures. Biotechnol Bioeng 94(2):373–382

    Article  CAS  Google Scholar 

  • Wong DC, Wong KT, Nissom PM, Heng CK, Yap MG (2006b) Targeting early apoptotic genes in batch and fed-batch CHO cell cultures. Biotechnol Bioeng 95(3):350–361

    Article  CAS  Google Scholar 

  • Wu SC (2009) RNA interference technology to improve recombinant protein production in Chinese hamster ovary cells. Biotechnol Adv 27(4):417–422

    Article  CAS  Google Scholar 

  • Wu SC, Hong WW, Liu JH (2008) Short hairpin RNA targeted to dihydrofolate reductase enhances the immunoglobulin G expression in gene-amplified stable Chinese hamster ovary cells. Vaccine 26(38):4969–4974

    Article  CAS  Google Scholar 

  • Xu C, Lu Y, Pan Z, Chu W, Luo X, Lin H, Xiao J, Shan H, Wang Z, Yang B (2007) The muscle-specific microRNAs miR-1 and miR-133 produce opposing effects on apoptosis by targeting HSP60, HSP70 and caspase-9 in cardiomyocytes. J Cell Sci 120(Pt 17):3045–3052

    Article  CAS  Google Scholar 

  • Yamane-Ohnuki N, Kinoshita S, Inoue-Urakubo M, Kusunoki M, Iida S, Nakano R, Wakitani M, Niwa R, Sakurada M, Uchida K, Shitara K, Satoh M (2004) Establishment of FUT8 knockout Chinese hamster ovary cells: an ideal host cell line for producing completely defucosylated antibodies with enhanced antibody-dependent cellular cytotoxicity. Biotechnol Bioeng 87(5):614–622

    Article  CAS  Google Scholar 

  • Yang SH, Cheng PH, Sullivan RT, Thomas JW, Chan AW (2008) Lentiviral integration preferences in transgenic mice. Genesis 46(12):711–718

    Article  CAS  Google Scholar 

  • Yeom KH, Lee Y, Han J, Suh MR, Kim VN (2006) Characterization of DGCR8/Pasha, the essential cofactor for Drosha in primary miRNA processing. Nucleic Acids Res 34(16):4622–4629

    Article  CAS  Google Scholar 

  • Zhang M, Koskie K, Ross JS, Kayser KJ, Caple MV (2010) Enhancing glycoprotein sialylation by targeted gene silencing in mammalian cells. Biotechnol Bioeng 105(6):1094–1105

    CAS  Google Scholar 

  • Zhang X, Lok SH, Kon OL (1998) Stable expression of human alpha-2, 6-sialyltransferase in Chinese hamster ovary cells: functional consequences for human erythropoietin expression and bioactivity. Biochim Biophys Acta 1425(3):441–452

    CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Cell Culture Technology, Faculty of Technology, Bielefeld University, Bielefeld, Germany

    Oliver Krämer, Sandra Klausing & Thomas Noll

  2. Center for Biotechnology (CeBiTec), Bielefeld University, Bielefeld, Germany

    Oliver Krämer, Sandra Klausing & Thomas Noll

  3. Bielefeld University, Universitaetsstrasse 25, 33615, Bielefeld, Germany

    Thomas Noll

Authors
  1. Oliver Krämer
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Sandra Klausing
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Thomas Noll
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Thomas Noll.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Krämer, O., Klausing, S. & Noll, T. Methods in mammalian cell line engineering: from random mutagenesis to sequence-specific approaches. Appl Microbiol Biotechnol 88, 425–436 (2010). https://doi.org/10.1007/s00253-010-2798-6

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00253-010-2798-6

Keywords

Search

Navigation