Long-term stability of β-galactosidase protein expression in the absence of selective growth conditions in transfected Chinese hamster ovary cell

Norha Saifudin, Nurina Anuar, Nazlina Ibrahim

Research output: Contribution to journalArticle

Abstract

In this study, the adequacy of β-galactosidase (β-gal) as marker for models that requires durable and high level gene expression in the absence of selective pressure was investigated. Chinese hamster ovary (CHO) cells were transfected with expression vector pcDNA4/HisMax-TOPO/lacZ containing lacZ and zeocin resistant genes. 26 recombinant CHO cell lines were established using lipid cationic transfection (TransFast™ Transfection Reagent) as DNA transfer method. 6 clones were productive in the expression of β-gal when grown in the presence of zeocin as the selective pressure. 2 sub-clones, TF8 (1) and TF9 (7) grown for 11 passages in nonselective medium which maintained high levels of protein expression with specific β-gal activity in the absence of zeocin were almost constant at 1.5704 and 4.3017 units β-gal activity/mg protein respectively. Specific growth rate of TF8 (1) and TF9 (7) in the absence of zeocin were approximately 0.638 and 0.656 day -1 respectively. The expression of β-gal does not affect the cell growth and the transfectants were stable population in terms of cell viability. Removal of zeocin from the media increases specific growth rate from a range of 24 to 52% and β-gal protein production in reference to specific activity increases from 128 to 320% with the capability to be expanded to larger volumes. In this study, we demonstrate transfected CHO cells with the ability to produce β-gal without the presence of zeocin for at least 11 passages.

Original languageEnglish
Pages (from-to)19187-19196
Number of pages10
JournalAfrican Journal of Biotechnology
Volume10
Issue number82
DOIs
Publication statusPublished - 19 Dec 2011

Fingerprint

Galactosidases
galactosidases
Protein Stability
Chinese hamsters
Cricetulus
Ovary
protein synthesis
transfection
specific growth rate
Growth
clones
transfer DNA
cells
cell viability
Transfection
cell growth
proteins
Clone Cells
cell lines
gene expression

Keywords

  • β-galactosidase
  • Chinese hamster ovary (CHO)
  • Clone stability
  • Selective pressure

ASJC Scopus subject areas

  • Biotechnology
  • Applied Microbiology and Biotechnology
  • Genetics
  • Molecular Biology
  • Agronomy and Crop Science

Cite this

@article{8997ae758b8546cbb207ed409c2aa976,
title = "Long-term stability of β-galactosidase protein expression in the absence of selective growth conditions in transfected Chinese hamster ovary cell",
abstract = "In this study, the adequacy of β-galactosidase (β-gal) as marker for models that requires durable and high level gene expression in the absence of selective pressure was investigated. Chinese hamster ovary (CHO) cells were transfected with expression vector pcDNA4/HisMax-TOPO/lacZ containing lacZ and zeocin resistant genes. 26 recombinant CHO cell lines were established using lipid cationic transfection (TransFast™ Transfection Reagent) as DNA transfer method. 6 clones were productive in the expression of β-gal when grown in the presence of zeocin as the selective pressure. 2 sub-clones, TF8 (1) and TF9 (7) grown for 11 passages in nonselective medium which maintained high levels of protein expression with specific β-gal activity in the absence of zeocin were almost constant at 1.5704 and 4.3017 units β-gal activity/mg protein respectively. Specific growth rate of TF8 (1) and TF9 (7) in the absence of zeocin were approximately 0.638 and 0.656 day -1 respectively. The expression of β-gal does not affect the cell growth and the transfectants were stable population in terms of cell viability. Removal of zeocin from the media increases specific growth rate from a range of 24 to 52{\%} and β-gal protein production in reference to specific activity increases from 128 to 320{\%} with the capability to be expanded to larger volumes. In this study, we demonstrate transfected CHO cells with the ability to produce β-gal without the presence of zeocin for at least 11 passages.",
keywords = "β-galactosidase, Chinese hamster ovary (CHO), Clone stability, Selective pressure",
author = "Norha Saifudin and Nurina Anuar and Nazlina Ibrahim",
year = "2011",
month = "12",
day = "19",
doi = "10.5897/AJB10.2220",
language = "English",
volume = "10",
pages = "19187--19196",
journal = "African Journal of Biotechnology",
issn = "1684-5315",
publisher = "Academic Journals",
number = "82",

}

TY - JOUR

T1 - Long-term stability of β-galactosidase protein expression in the absence of selective growth conditions in transfected Chinese hamster ovary cell

AU - Saifudin, Norha

AU - Anuar, Nurina

AU - Ibrahim, Nazlina

PY - 2011/12/19

Y1 - 2011/12/19

N2 - In this study, the adequacy of β-galactosidase (β-gal) as marker for models that requires durable and high level gene expression in the absence of selective pressure was investigated. Chinese hamster ovary (CHO) cells were transfected with expression vector pcDNA4/HisMax-TOPO/lacZ containing lacZ and zeocin resistant genes. 26 recombinant CHO cell lines were established using lipid cationic transfection (TransFast™ Transfection Reagent) as DNA transfer method. 6 clones were productive in the expression of β-gal when grown in the presence of zeocin as the selective pressure. 2 sub-clones, TF8 (1) and TF9 (7) grown for 11 passages in nonselective medium which maintained high levels of protein expression with specific β-gal activity in the absence of zeocin were almost constant at 1.5704 and 4.3017 units β-gal activity/mg protein respectively. Specific growth rate of TF8 (1) and TF9 (7) in the absence of zeocin were approximately 0.638 and 0.656 day -1 respectively. The expression of β-gal does not affect the cell growth and the transfectants were stable population in terms of cell viability. Removal of zeocin from the media increases specific growth rate from a range of 24 to 52% and β-gal protein production in reference to specific activity increases from 128 to 320% with the capability to be expanded to larger volumes. In this study, we demonstrate transfected CHO cells with the ability to produce β-gal without the presence of zeocin for at least 11 passages.

AB - In this study, the adequacy of β-galactosidase (β-gal) as marker for models that requires durable and high level gene expression in the absence of selective pressure was investigated. Chinese hamster ovary (CHO) cells were transfected with expression vector pcDNA4/HisMax-TOPO/lacZ containing lacZ and zeocin resistant genes. 26 recombinant CHO cell lines were established using lipid cationic transfection (TransFast™ Transfection Reagent) as DNA transfer method. 6 clones were productive in the expression of β-gal when grown in the presence of zeocin as the selective pressure. 2 sub-clones, TF8 (1) and TF9 (7) grown for 11 passages in nonselective medium which maintained high levels of protein expression with specific β-gal activity in the absence of zeocin were almost constant at 1.5704 and 4.3017 units β-gal activity/mg protein respectively. Specific growth rate of TF8 (1) and TF9 (7) in the absence of zeocin were approximately 0.638 and 0.656 day -1 respectively. The expression of β-gal does not affect the cell growth and the transfectants were stable population in terms of cell viability. Removal of zeocin from the media increases specific growth rate from a range of 24 to 52% and β-gal protein production in reference to specific activity increases from 128 to 320% with the capability to be expanded to larger volumes. In this study, we demonstrate transfected CHO cells with the ability to produce β-gal without the presence of zeocin for at least 11 passages.

KW - β-galactosidase

KW - Chinese hamster ovary (CHO)

KW - Clone stability

KW - Selective pressure

UR - http://www.scopus.com/inward/record.url?scp=84855759739&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84855759739&partnerID=8YFLogxK

U2 - 10.5897/AJB10.2220

DO - 10.5897/AJB10.2220

M3 - Article

VL - 10

SP - 19187

EP - 19196

JO - African Journal of Biotechnology

JF - African Journal of Biotechnology

SN - 1684-5315

IS - 82

ER -