The Construction Of The Plasmid Vector Of Human Parathyroid Hormone Gene

Parathyroid hormone (PTH) is a dominant homeostatic regulator of blood calcium and phosphate. Parathyroid damage, tumors or genetic factors can all result in the hyposecretion of PTH and the decrease of the biologic activity of PTH, which is called hypothyroidism. Recently, the success of clone and expression of related gene make the gene therapy for hypothyroidism possible. We designed a pair of promoters basing on nucieotide sequence of PTH gene reported, introducing two restriction enzyme sites (EcoR I ,Xba I )into the promoters. In this report we acquired the gene of PTH from the total RNA from human fetal parathyroid tissues by RT-PCR, Furthermore, we cloned the fragment into the plasmid PCR3.I. The recombinant plasmid containing inserted fragment was acquired after transformed by Escherichia coil JMIO9. Then , the positive recombinant plasmids were determined and sequenced. we confirm that we acquired the expression recombinant by two channels.The one is that we obtained positive fragment (270bp) by the PCR amplification of the recombinant plasmid. The other is two fragments could be seen on gel (270bp.5.Okb) after digested by EcoRL XbaI. We Conclude that the goal gene fragment has been cloned into PCR3. 1 vector. The sequencing and sequence analysis indicated that the inserted fragment was PTH gene. The achievement of this study established the foundation for the further therapy of hypothyroidism.