Sub-cloning of alpha subunit of anterior pituitary glycoprotein hormone into a mammalian expression vector
Author(s):
Abstract:
Introduction
Anterior pituitary glycoprotein hormones include thyroid stimulating hormone, lutinizing hormone, follicule stimulating hormone, and gonadotropine hormone. Each of them contains alpha and beta subunits. The alpha subunit gene is the same in all of these hormones and contains 4 exones and 3 intrones. The beta subunit is responsible for specific function of each hormone. The aim of this study was to clone alpha chain cDNA of glycoprotein hormones in a proper vector for eukaryotic system. Material And Methods
To clone cDNA, alpha subunit of glycoprotein hormones was amplified by using one pair primers and T.vector as template and cloned in Not I and Bam HI sites of pcDNA3.1 plasmid. The recombinant plasmid transformed to E.coli Top10F΄ cell and colonies that contain plasmid were selected by Colony PCR. The accuracy of extracted plasmid of these clones was approved by enzyme digestion and sequencing. Results
Enzyme analysis showed that pcDNA3.1-F351 a had correct structure and sequencing confirmed by 100% homology of the gene with reported alpha gene in Gene Bank. Conclusion
Because of its proper structure, this plasmid is able to transform to Eukaryotic system and translation.Language:
Persian
Published:
Koomesh, Volume:13 Issue: 2, 2011
Page:
143
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