Codon Optimization, Cloning and Expression of the Human Leukemia Inhibitory Factor (hLIF) in E. coli

Authors

  • Ali Khodadadi Cancer Research Center, Ahvaz Jundishapur University of Medical School, Ahvaz, IR Iran
  • Amir Jalali Research Center of Toxicology, Ahvaz Jundishapur University of Medical School, Ahvaz, IR Iran
  • Hamid Galehdari Department of Genetics, Shahid Chamran University, Ahvaz, IR Iran
  • Mohammad Shafeei Department of Genetics, Shahid Chamran University, Ahvaz, IR Iran
  • Nahid Shahbazian Department of Gynecology, Ahvaz Jundishapur University of medical School, Ahvaz, IR Iran
  • Parichehr Darabi Department of Genetics, Shahid Chamran University, Ahvaz, IR Iran
  • Saeed Reza Khatami Department of Genetics, Shahid Chamran University, Ahvaz, IR Iran
Abstract:

Background: Leukemia inhibitor factor (LIF) is a very important pleiotropic cytokine which belongs to interleukin-6 (IL-6) family. LIF exerts multiple effects on different types of cells and tissues with numerous regulatory effects in vivo and in vitro. It is a lymphoid factor, which performs a number of activities including cholinergic neuron differentia‌tion, control of stem cell pluripotency, bone and fat metabolism, and is important for embryo implantation and promoting megakaryocytes production in vivo. Human LIF is a potential therapeutic candidate for some diseases such as multiplesclerosis (MS). Objectives: Because of aforementioned applications of the LIF protein in biological sys‌tems, the LIF gene has been cloned in various species. In this study a useful novel method was used to clone an optimized LIF sequence. Materials and Methods: In this study, the optimized cDNA form of human leukemia inhibitory factorwas cloned into the pET-28a (+) expression vector under control of T7lacpromoter using BamHI and XbaI restriction enzymes. The recombinant vector was transformed into Escherichia coli strain BL21 (DE3). The cloned hLIF cDNA was expressed as a fusion protein with His-tag. Cloning of hLIF cDNA was confirmed by digestion and DNA sequencing. Appropriate expression of recombinant hLIF was examined by SDS-PAGE af‌ter induction with isopropylthio-β-galactoside (IPTG). Results: The results confirmed the expression of the 19.7 kDa rhLIF protein in the bacte‌rial expression system. Conclusions: We could show that codon optimization might increase the production of recombinant hLIF in the E. coli. This result is useful in similar cases, in which the level of expressed gene is critical.

Upgrade to premium to download articles

Sign up to access the full text

Already have an account?login

similar resources

codon optimization, cloning and expression of the human leukemia inhibitory factor (hlif) in e. coli

background: leukemia inhibitor factor (lif) is a very important pleiotropic cytokine which belongs to interleukin-6 (il-6) family. lif exerts multiple effects on different types of cells and tissues with numerous regulatory effects in vivo and in vitro. it is a lymphoid factor, which performs a number of activities including cholinergic neuron differentia­tion, control of stem cell pluripotency...

full text

Cloning, Codon Optimization, and Expression of Yersinia intermedia Phytase Gene in E. coli

Background: Phytate is an anti-nutritional factor in plants, which catches the most phosphorus contents and some vital minerals. Therefore, Phytase is added mainly as an additive to the monogastric animals’ foods to hydrolyze phytate and increase absorption of phosphorus. Objectives: Y. intermedia phytase is a new phytase with special characteristics such as high specific activity, pH stabilit...

full text

CLONING AND EXPRESSION OF A HUMAN INTERFERON a2 GENE IN E. COLI

The plasmid pALCA1SIFN containing cDNA that encodes the human interferon a-2b was obtained from the ATCC(no. 531667). In this system the expression of the gene is under the control of an alcA promoter. alcA p is a specific promoter for expression of different genes in Aspergillusfilamentous. In this plasmid the coding region of IFN?-2b is preceded by the coding region of a synthetic signal ...

full text

cloning, expression, and functional characterization of in-house prepared human leukemia inhibitory factor

objective: leukemia inhibitory factor (lif) plays important roles in cellular proliferation, growth promotion and differentiation of various types of target cells. in addition, lif influences bone metabolism, cachexia, neural development, embryogenesis and inflammation. human lif (hlif) is an essential growth factor for the maintenance of mouse embryonic stem cells (escs) and induced pluripoten...

full text

Cloning and Expression of Human Gamma-Interferon cDNA in E. coli

Prior to the production of human gamma interferon using recombinant DNA technology, it had been producedmainly upon mitogenic induction of lymphocytes in very low amounts, which evidently hamperedits characterization and its medical applications. The recombinant gamma interferons produced in largerquantities in prokaryotic systems retain their biological activities, and can be...

full text

RT-PCR MEDIATED CLONING OF HUMAN GROWTH HORMONE GENE AND I TS EXPRESSION IN E. coli

Human growth hormone (hGH) genomic sequence containing 5 exons and 4 introns was cloned in pcDNA-3 and the constructed plasmid was subsequently used for transfection ofNlli-3T3 cell line using lipofection technique. Expression of hGH in stably transfected cells was assayed using ELISA. Total RNA was extracted from transfected cells and hGH cDNA was amplified by RT-PCR using specific primers...

full text

My Resources

Save resource for easier access later

Save to my library Already added to my library

{@ msg_add @}


Journal title

volume 11  issue 1

pages  47- 53

publication date 2013-01-01

By following a journal you will be notified via email when a new issue of this journal is published.

Hosted on Doprax cloud platform doprax.com

copyright © 2015-2023