Volume 13, number 4
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Evaluation of Interferon-Beta  Protein Expression in HEK293T Cell-Line Transfected by Recombinant Construction, Pbud.Ifnβ-1a

Raheleh Norouzi1, Zohreh Hojati2  and Zahra Badr1

1Biology Department, Genetic section, Faculty of Sciences, University of Isfahan, Isfahan, Iran.

2Isfahan University, Department of Biology, Genetics section, Faculty of Sciences, University of Isfahan, Isfahan, Iran.

DOI : http://dx.doi.org/10.13005/bbra/2392

ABSTRACT: Cytokines  are  a  group  of  signaling  glycoproteins  that  are  expressed  in  respond  to  stimulus  factors  and  antigens.  IFNβ  is  an  important  cytokine  that  play  role  in  immune  processes,  inflammation  and  host  defense  against  external  factors.  In  this  study,  the  expression  level  of  IFNβ  protein  in  HEK293T  cell  line  transfected  by  recombinant  construction  pBud.IFNβ  examine  via  Dot  blot  and  Elisa  protein  tests.  Sequence  of  IFNβ  gene  with  using  of  primers  containing  Kozak  conserved  sequence  and  enzyme  restriction  site  of  Kpnl  and  BglII  amplified  from  pSVMdhfr  vector.  After  treatment  by  above  mentioned  enzymes,  IFNβ  gene  cloned  in  linearized  pBud.CE4.1.  Structure  of  recombinant  vector  were  examined  via  RFLP,  Colony  PCR  and  sequencing  and  finally  transformed  into  competent  Ecoli.TOP10  bacteria  and  next  transfected  to  HEK293T  cells.  The  results  of  protein  tests  indicate  that  recombinant  IFNβ  gene  expressed  successfully  in  HEK293T  cell  line  under  eEf1a  promoter  of  selected  vector.  Proteins  produced  in  prokaryotic  systems  are  lacking  glycosylation  and  thus  have  different  physicochemical  properties  in  comparison  with  normal  protein  production  in  the  body.  This  causes  side  effects  in  the  patient  so  the  production  of  IFNβ  protein  in  human  HEK293T  cells  and  high  expression  level  of  this  protein  is  the  benefits  of  this  research.

KEYWORDS: Recombinant  interferon beta;  HEK293T  cell  line; pBud.CE4.1;  dot  blot;  ELISA

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Norouzi R, Hojati Z, Badr Z. Evaluation of Interferon-Beta Protein Expression in HEK293T Cell-Line Transfected by Recombinant Construction, Pbud.Ifnβ-1a. Biosci Biotech Res Asia 2016;13(4).
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Norouzi R, Hojati Z, Badr Z. Evaluation of Interferon-Beta Protein Expression in HEK293T Cell-Line Transfected by Recombinant Construction, Pbud.Ifnβ-1a. Biosci Biotech Res Asia 2016;13(4). Available from: https://www.biotech-asia.org/?p=16788

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