Calcium phosphate transfection generates mammalian recombinant cell lines with higher specific productivity than polyfection.

Abstract:

:Transfection with polyethylenimine (PEI) was evaluated as a method for the generation of recombinant Chinese hamster ovary (CHO DG44) cell lines by direct comparison with calcium phosphate-DNA coprecipitation (CaPO4) using both green fluorescent protein (GFP) and a monoclonal antibody as reporter proteins. Following transfection with a GFP expression vector, the proportion of GFP-positive cells as determined by flow cytometry was fourfold higher for the PEI transfection as compared to the CaPO4 transfection. However, the mean level of transient GFP expression for the cells with the highest level of fluorescence was twofold greater for the CaPO4 transfection. Fluorescence in situ hybridization on metaphase chromosomes from pools of cells grown under selective pressure demonstrated that plasmid integration always occurred at a single site regardless of the transfection method. Importantly, the copy number of integrated plasmids was measurably higher in cells transfected with CaPO4. The efficiency of recombinant cell line recovery under selective pressure was fivefold higher following PEI transfection, but the average specific productivity of a recombinant antibody was about twofold higher for the CaPO4-derived cell lines. Nevertheless, no difference between the two transfection methods was observed in terms of the stability of protein production. These results demonstrated the feasibility of generating recombinant CHO-derived cell lines by PEI transfection. However, this method appeared inferior to CaPO4 transfection with regard to the specific productivity of the recovered cell lines.

journal_name

Biotechnol Bioeng

authors

Chenuet S,Martinet D,Besuchet-Schmutz N,Wicht M,Jaccard N,Bon AC,Derouazi M,Hacker DL,Beckmann JS,Wurm FM

doi

10.1002/bit.21972

subject

Has Abstract

pub_date

2008-12-01 00:00:00

pages

937-45

issue

5

eissn

0006-3592

issn

1097-0290

journal_volume

101

pub_type

杂志文章
  • On the organic solvent and thermostability of the biocatalytic redox system of Rhodococcus ruber DSM 44541.

    abstract::The sec-alcohol dehydrogenase activity of whole cells of Rhodococcus ruber DSM 44541 has been employed as an efficient biocatalytic redox system due to the use of acetone and 2-propanol at elevated concentrations for cofactor regeneration in the oxidation and reduction mode, respectively, and external addition of NADH...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.10540

    authors: Stampfer W,Kosjek B,Kroutil W,Faber K

    更新日期:2003-03-30 00:00:00

  • Affinity purification of plasmid DNA directly from crude bacterial cell lysates.

    abstract::We have shown previously that a sequence-specific DNA-binding protein based on the Lac repressor protein can isolate pre-purified DNA efficiently from simple buffer solution but our attempts to purify plasmids directly from crude starting materials were disappointing with impractically low DNA yields. We have optimize...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.21492

    authors: Darby RA,Forde GM,Slater NK,Hine AV

    更新日期:2007-12-01 00:00:00

  • Effect of temperature on nucleotide pools and monoclonal antibody production in a mouse hybridoma.

    abstract::The specific monoclonal antibody productivity (q(Mab)) of a murine hybridoma (CC9C10) increased with incubation temperature in the range 33 degrees C to 39 degrees C. q(Mab) was constant at each temperature and was independent of the phase of culture. The q(Mab) increased 97% at 39 degrees C and decreased by 21% at 33...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260441011

    authors: Barnabé N,Butler M

    更新日期:1994-11-20 00:00:00

  • Kinetic analysis of bacterial bioluminescence.

    abstract::Bioluminescence from the lux-based bacterial reporter Pseudomonas fluorescens HK44 was experimentally investigated under growth substrate-rich and limiting conditions in batch, continuous stirred tank (CSTR), and turbidostat reactors. A mechanistically based, mathematical model was developed to describe bioluminescenc...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.10475

    authors: Kelly CJ,Hsiung CJ,Lajoie CA

    更新日期:2003-02-05 00:00:00

  • Three-dimensional biofilm model with individual cells and continuum EPS matrix.

    abstract::An innovative type of biofilm model is derived by combining an individual description of microbial particles with a continuum representation of the biofilm matrix. This hybrid model retains the advantages of each approach, while providing a more realistic description of the temporal development of biofilm structure in...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.20917

    authors: Alpkvist E,Picioreanu C,van Loosdrecht MC,Heyden A

    更新日期:2006-08-05 00:00:00

  • H2 production by photofermentation in an innovative plate-type photobioreactor with meandering channels.

    abstract::Hydrogen production by Rhodobacter capsulatus is an anaerobic, photobiological process requiring specific mixing conditions. In this study, an innovative design of a photobioreactor is proposed. The design is based on a plate-type photobioreactor with an interconnected meandering channel to allow culture mixing and H2...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.27656

    authors: Turon V,Ollivier S,Cwicklinski G,Willison JC,Anxionnaz-Minvielle Z

    更新日期:2020-12-15 00:00:00

  • Effects of culture conditions on N-glycolylneuraminic acid (Neu5Gc) content of a recombinant fusion protein produced in CHO cells.

    abstract::CHO cells express glycoproteins containing both the N-acetylneuraminic acid (Neu5Ac) and minor amounts of the N-glycolylneuraminic acid (Neu5Gc) forms of sialic acid. As Neu5Gc is not expressed in humans and can be recognized as a foreign epitope, there is the potential for immunogenicity issues for glycoprotein thera...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.22644

    authors: Borys MC,Dalal NG,Abu-Absi NR,Khattak SF,Jing Y,Xing Z,Li ZJ

    更新日期:2010-04-15 00:00:00

  • Trichloroethene and cis-1,2-dichloroethene concentration-dependent toxicity model simulates anaerobic dechlorination at high concentrations. II: continuous flow and attached growth reactors.

    abstract::A model that was used to describe toxicity from high concentrations of chlorinated aliphatic hydrocarbons (CAHs) on reductively dechlorinating cultures in batch reactors (Sabalowsky and Semprini (in press)) was extended here to simulate observations in continuous flow suspended and attached growth reactors. The reduct...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.22822

    authors: Sabalowsky AR,Semprini L

    更新日期:2010-10-15 00:00:00

  • Impact of clarification strategy on chromatographic separations: Pre-processing of cell homogenates.

    abstract::This research focused on how the extent and type of primary solid-liquid separation can affect the performance of guard filtration and chromatography, in this instance hydrophobic interaction chromatography. The system used in the study was yeast (Saccharomyces cerevisiae) with the target molecule being an intracellul...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.21823

    authors: Bracewell DG,Boychyn M,Baldascini H,Storey SA,Bulmer M,More J,Hoare M

    更新日期:2008-08-01 00:00:00

  • Analysis of continuous cultures of recombinant methylotrophs.

    abstract::Static and dynamic characteristics of continuous cultures of recombinant methylotrophs, which are designed to improve the selectivity of plasmid-bearing cells and the plasmid stability, are investigated in detail. Operational regions in which coexistence (survival of plasmid-bearing and plasmid-free cells) operation i...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260290802

    authors: Parulekar SJ,Chang YK,Modak JM,Lim HC

    更新日期:1987-06-01 00:00:00

  • Optimized delignification of wood-derived lignocellulosics for improved enzymatic hydrolysis.

    abstract::One of the major bottlenecks in the bioconversion of lignocelluosic feedstocks to liquid ethanol is the recalcitrance of residue following pretreatment, specifically softwood derived residues. Peroxide delignification has previously been shown to effectively aid in the removal of condensed lignaceous moieties from sub...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.22768

    authors: Cullis IF,Mansfield SD

    更新日期:2010-08-15 00:00:00

  • Improvement of product yields by temperature-shifting of Escherichia coli cultures containing plasmid pOU140.

    abstract::Temperature shifting was investigated as a means of improving cloned-gene product yields form a recombinant Escherichia coli containing the temperature-sensitive plasmid, pOU 140. In a series of shaker flask fermentations recombinant cells were thermally induced for different time periods. The growth, stability, and p...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260290416

    authors: Dipasquantonio VM,Betenbaugh MJ,Dhurjati P

    更新日期:1987-03-01 00:00:00

  • Enhanced purification of plasmid DNA isoforms by exploiting ionic strength effects during ultrafiltration.

    abstract::The solution structure of plasmid DNA is known to be a strong function of solution conditions due to intramolecular electrostatic interactions between the charged phosphate groups along the DNA backbone. The objective of this work was to determine whether it was possible to enhance the use of ultrafiltration for separ...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.25836

    authors: Li Y,Currie D,Zydney AL

    更新日期:2016-04-01 00:00:00

  • Comparison of microbial and transient expression (tobacco plants and plant-cell packs) for the production and purification of the anticancer mistletoe lectin viscumin.

    abstract::Cancer is the leading cause of death in industrialized countries. Cancer therapy often involves monoclonal antibodies or small-molecule drugs, but carbohydrate-binding lectins such as mistletoe (Viscum album) viscumin offer a potential alternative treatment strategy. Viscumin is toxic in mammalian cells, ruling them o...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.27076

    authors: Gengenbach BB,Keil LL,Opdensteinen P,Müschen CR,Melmer G,Lentzen H,Bührmann J,Buyel JF

    更新日期:2019-09-01 00:00:00

  • Enhanced d-lactic acid production by recombinant Saccharomyces cerevisiae following optimization of the global metabolic pathway.

    abstract::Utilization of renewable feedstocks for the production of bio-based chemicals such as d-lactic acid by engineering metabolic pathways in the yeast Saccharomyces cerevisiae has recently become an attractive option. In this study, to realize efficient d-lactic acid production by S. cerevisiae, the expression of 12 glyco...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.26330

    authors: Yamada R,Wakita K,Mitsui R,Ogino H

    更新日期:2017-09-01 00:00:00

  • Estimation of the true growth yield and maintenance coefficient for yoghurt cultures.

    abstract::Point and 95% confidence interval estimates of the true growth yield and maintenance coefficient are reported for four pure cultures andtwo mixed cultures of Lactobacillus bulgaricus and Streptococcus thermophilus grown anaerobically on 3% nonfat dry milk using batch followed by fed batch culture. An improved method o...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260280619

    authors: Oner MD,Erickson LE,Yang SS

    更新日期:1986-06-01 00:00:00

  • Toward an enzyme-based oxygen scavenging laminate. Influence of industrial lamination conditions on the performance of glucose oxidase.

    abstract::The laminate consisted of several polymer layers, aluminium, and one cellulose-based layer containing the active enzymatic system (e.g., glucose oxidase, catalase, glucose, and CaCO3). During the industrial lamination process, the enzyme layer was exposed to three temperature spikes up to 325 degrees C without signifi...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.10266

    authors: Andersson M,Andersson T,Adlercreutz P,Nielsen T,Hörnsten EG

    更新日期:2002-07-05 00:00:00

  • Cellular reporoduction and extracellular polymer formation by Pseudomonas aeruginosa in continuous culture.

    abstract::The kinetics of cellular reproduction and the rate and extent of synthesis of extracellular polymeric substances (EPS) were investigated for P. aeruginosa growing in glucose-limited chemostats. mu(max) and K(s) estimates of 0.4 h(-1) and 2 mg glucose C/L, respectively, at 25 degrees C were obtained for this bacterium....

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260261203

    authors: Robinson JA,Trulear MG,Characklis WG

    更新日期:1984-12-01 00:00:00

  • Entrapment of concanavalin A-glycoenzyme complexes in calcium alginate gels.

    abstract::Glucose oxidase, invertase, and amyloglucosidase were entrapped in calcium alginate gels as concanavalin A complexes in order to prevent the leaching out of the enzymes from the porous matrix. The free as well as the gel-entrapped concanavalin A-glycoenzyme complexes exhibited a relatively high effectiveness factor, e...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260270803

    authors: Husain Q,Iqbal J,Saleemuddin M

    更新日期:1985-08-01 00:00:00

  • Effect of pH change on the performance and microbial community of enhanced biological phosphate removal process.

    abstract::An acetate-rich wastewater, containing 170 mg/L of total organic carbon (TOC), 13 mg/L of N, and 15 mg/L of P, was treated using the enhanced biological phosphate removal (EBPR) process operated in a sequencing batch reactor. A slight change of pH of the mixed liquor from 7.0 to 6.5 led to a complete loss of phosphate...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.20589

    authors: Zhang T,Liu Y,Fang HH

    更新日期:2005-10-20 00:00:00

  • A thermodynamic model to predict Phanerochaete chrysosporium INA-12 adhesion to various solid carriers in relation to lignin peroxidase production.

    abstract::A thermodynamic model was used in this study to predict the adhesion of Phanerochaete chrysosporium INA-12 as conidiospores or mycelium to various solid carriers. Theoretical predictions were closely reflected by experimental results. Amount of immobilized mycelium was higher for hydrophobic (polypropylene and polyure...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260350505

    authors: Asther M,Bellon-Fontaine MN,Capdevila C,Corrieu G

    更新日期:1990-03-05 00:00:00

  • Engineering CHO cells to overexpress a secreted reporter protein upon induction from mouse mammary tumor virus promoter.

    abstract::The mouse mammary tumor virus (MMTV) promoter is induced by the addition of a glucocorticoid hormone or analog such as dexamethasone. The hormone binds to its specific transcription factor, glucocorticoid receptor (GR), and the activated complex then binds to the glucocorticoid response elements (GREs) in the enhancer...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:

    authors: James RI,Elton JP,Todd P,Kompala DS

    更新日期:2000-01-20 00:00:00

  • Macrokinetic and quantitative microbial investigation on a bench-scale biofilter treating styrene-polluted gaseous streams.

    abstract::We performed a macrokinetic and quantitative microbial investigation of a continuously operating bench-scale biofilter treating styrene-polluted gases. The device was filled with a mixture of peat and glass beads as packing medium and inoculated with the styrene-oxidizing strain, Rhodococcus rhodochrous AL NCIMB 13259...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.10640

    authors: Zilli M,Converti A,Di Felice R

    更新日期:2003-07-05 00:00:00

  • Dependence of biological treatment rate on species composition in activated sludge or biofilm. II: From models to theory.

    abstract::Traditional biological treatment models are "deduced" from formal chemical kinetics or dynamics of pure microorganism cultures growth. The best formal models give reasonable approximations of the biological treatment model with an ecosystem adaptation (ESA model). The model presented here explains some features of the...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260250610

    authors: Vavilin VA

    更新日期:1983-06-01 00:00:00

  • Fiber alignment and coculture with fibroblasts improves the differentiated phenotype of murine embryonic stem cell-derived cardiomyocytes for cardiac tissue engineering.

    abstract::Embryonic stem cells (ESCs) are an important source of cardiomyocytes for regenerating injured myocardium. The successful use of ESC-derived cardiomyocytes in cardiac tissue engineering requires an understanding of the important scaffold properties and culture conditions to promote cell attachment, differentiation, or...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.23353

    authors: Parrag IC,Zandstra PW,Woodhouse KA

    更新日期:2012-03-01 00:00:00

  • Optimization of batch processes involving simultaneous enzymatic and microbial reactions.

    abstract::Two general models for batch simultaneous enzymatic and microbial reaction (SEMR) processes are presented, the second derived from and simpler than the first and accounting for enzyme denaturation. Using the second model and parameter values from the literature, simulation was used to examine a range of enzyme additio...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260371114

    authors: Asenjo JA,Sun WH,Spencer JL

    更新日期:1991-05-01 00:00:00

  • FIA for on-line monitoring of important lactic acid fermentation variables.

    abstract::An automated system with semi on-line monitoring of glucose, lactic acid, protein, and optical density during lactic acid fermentations, is set up to study the dynamics of lactic acid bacteria. The analyzers for glucose, lactic acid, and protein are based on flow injection analysis (FIA). The system consists of a labo...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260330908

    authors: Nielsen J,Nikolajsen K,Villadsen J

    更新日期:1989-04-05 00:00:00

  • Affinity-based reversed micellar protein extraction: II. effect of cosurfactant tail length.

    abstract::The selectivity of protein extraction by reversed micellar solutions can be improved by the addition of affinity cosurfactants bearing ligands which bind strongly to the target protein. The interactions between cosurfactant and protein, as well as the interfacial activity of both the free cosurfactant and the protein-...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260421011

    authors: Kelley BD,Wang DI,Hatton TA

    更新日期:1993-11-20 00:00:00

  • Application of empirical design methodologies to the study of the influence of reaction conditions and N-alpha-protecting group structure on the enzymatic X-Phe-Leu-NH(2) dipeptide synthesis in buffer/dimethylformamide solvents systems.

    abstract::The influence of five different N-terminal protecting groups (For, Ac, Boc, Z, and Fmoc) and reaction conditions (temperature and dimethylformamide content) on the alpha-chymotrypsin-catalyzed synthesis of the dipeptide derivative X-Phe-Leu-NH(2) was studied. Groups such as For, Ac, Boc, and Z always rendered good pep...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.260390509

    authors: Calvet S,Clapés P,Vigo JP,Xaus N,Jorba X,Mas RM,Torres JL,Valencia G,Serralheiro ML,Cabral JM,Empis JM

    更新日期:1992-03-05 00:00:00

  • A novel sugar analog enhances sialic acid production and biotherapeutic sialylation in CHO cells.

    abstract::A desirable feature of many therapeutic glycoprotein production processes is to maximize the final sialic acid content. In this study, the effect of applying a novel chemical analog of the sialic acid precursor N-acetylmannosamine (ManNAc) on the sialic acid content of cellular proteins and a model recombinant glycopr...

    journal_title:Biotechnology and bioengineering

    pub_type: 杂志文章

    doi:10.1002/bit.26291

    authors: Yin B,Wang Q,Chung CY,Bhattacharya R,Ren X,Tang J,Yarema KJ,Betenbaugh MJ

    更新日期:2017-08-01 00:00:00