en 30090207 <p>Background: Biosynthetic teriparatide (1-34) (TPD) is a N-terminally truncated version of human parathyroid hormone (hPTH). The recombinant form of this polypeptide has been expressed in <em>Escherichia coli (E. coli)</em> and approved as the first anabolic treatment of osteoporosis in the EU and the USA. Feasibility of expression and secretion of a tag- fused form of TPD into <em>Bacillus subtilis (B. subtilis)</em> was examined due to several advantages of <em>B. subtilis </em>over <em>E. coli </em>in production of recombinant proteins with pharmacological activities.<br /> Methods: A codon optimized gene containing TPD open reading frame carrying enterokinase site in its upstream was fully synthesized. According to our cloning scheme, this synthetic polynucleotide was used as a template for PCR amplification using engineered primers in such a way that a polyhistidin tag was added in frame to the upstream of the amplicon as well as two restriction sites at its ends. The resulted amplicon, a cassette containing His-tag, enterokinase site and TPD, from 5&rsquo; to 3&rsquo;, was cloned into pTZ57R/T vector and subjected to sequencing.The cassette was then subcloned into pHT43 shuttle vector and transformed into <em>B. subtilis</em>. Expression of target protein was analyzed by SDS-PAGE and western blotting upon induction by IPTG.<br /> Results: The accuracy of construction of pHT43-TPD was confirmed by sequencing and restriction map analyses. SDS-PAGE and western blotting results showed that the recombinant fusion form of hPTH was successfully expressed and secreted into cytoplasm and extracellular medium.<br /> Conclusion: TPD may be successfully expressed and secreted in <em>B. subtilis</em>; however, optimization of expression conditions is required for enhancing target protein yield.</p> Bacillus subtilis, hPTH, Teriparatide 147 151 https://www.ajmb.org/En/Article.aspx?id=319 https://www.ajmb.org/PDF/En/FullText/319.pdf MahdiKarimiDepartment of Molecular Genetics, Research Centre for Biosciences and Biotechnology, Malek-Ashtar University of Technology, Tehran, Iran1284FaridaBehzadianDepartment of Molecular Genetics, Research Centre for Biosciences and Biotechnology, Malek-Ashtar University of Technology, Tehran, Iran1285HamidehRouhaninejadDepartment of Molecular Genetics, Research Centre for Biosciences and Biotechnology, Malek-Ashtar University of Technology, Tehran, Iran1286SanazYariDepartment of Molecular Genetics, Research Centre for Biosciences and Biotechnology, Malek-Ashtar University of Technology, Tehran, Iran1287