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Construction of pcDNA3.1-FSHβ expression vector in order to transfer FSHβ gene into a mammalian cellline
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Reza نصر   , Mohammad reza اکبری عیدگاهی , Azar sadat رضاپور جوارشک , Ahmad reza بندگی , Saeid ولی زاده |
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Abstract: (11624 Views) |
| Introduction: Follicle stimulating hormone (FSH) is one of the glycoprotein hormones of
pituitary gland that consists of two subunits, alpha and beta. Beta subunit has
111 amino acids and a signal peptide, which is consisted of 18 amino acids and
it is responsible for biological activity of the hormone. The aim of this study
was to construct a pcDNA3.1FSHβ expression vector in order to transfer FSHβ gene
into a mammalian cell line. Materials and Methods: To sub-cloning of beta chain from T.vector, a pair of primer was designed
that they had a site for EcoRI and HindIII in addition of the start and stop
site of the beta chain gene. Amplified beta chain was cloned in pcDNA3.1 at the
site of the mentioned enzymes anvd the recombinant plasmid was transformed into
E.coli cell. The resulting colonies were checked by PCR re-amplification. The
plasmid was purified from some positive colonies. The accuracy of purified
plasmids were confirmed by both enzyme digestion and sequencing. Results: Enzyme analysis and sequencing demonstrated that pcDNA3.1-F390β had both
correct construction and gene sequence which had an exact correlation with
reported FSHβ gene in GenBank. Conclusion: This recombinant plasmid structurally is correct and is proper for transmitting
into mammalian cell culture. |
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| Keywords: Follicle stimulating hormone, β-subunit, Cloning, Mammalian cell line |
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Full-Text [PDF 342 kb]
(2635 Downloads)
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Type of Study: Research |
Subject:
General
Received: 2012/02/21 | Accepted: 2013/09/4 | Published: 2013/09/4
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