Human thyrotropin receptor extracellular and eukaryotic gene expression.docVIP

 PAGE \* MERGEFORMAT 16 Human thyrotropin receptor extracellular and eukaryotic gene expression Of: Li Xueping, Shibing silver, Wu Min, Wu Liping, Liu Shaanxi [Abstract] Objective To clone the human thyrotropin receptor extracellular domain gene, the recombinant eukaryotic expression plasmid, obtained with immunological activity of purified recombinant protein. Methods RT PCR from human thyroid tissue by extracellular TSHR cDNA, inserted eukaryotic expression vector pcDNA3.1 (+), transfected CHO cells, RT PCR identification of gene transcription, by immunocytochemistry staining and Western Blot Identification of protein. The results confirmed by sequencing and screened the recombinant TSHR exists in the correct sequence extracellular region gene expression of protein immunoreactivity. Conclusion cloned human thyrotropin receptor gene and the extracellular eukaryotic expression vector, and its successful expression of eukaryotic cells. [Keywords:] autoimmune thyroid disease; human thyrotropin receptor; gene recombination; eukaryotic expression ABSTRACT: Objective To clone and construct the plasmid containing human thyroid stimulating hormone receptor (TSHR) gene ectodomain, and then identify the immunoreactivity of the purified recombinant protein. Methods TSHR total RNA was extracted from human thyroid, and cDNA was obtained with RT PCR technique . Human TSHR ectodomain gene (hETSHR) was cloned into pcDNA3.1 (+) vector. The recombined construct was transfected into CHO cells by Lipofectin 2000. The transcript mRNA was detected by RT PCR, and protein immunoreactivity was assayed by TSHR antibody with immunocytochemistry staining and Western blot. Results DNA sequencing results showed that the recombinant of human thyrotropin receptor ectodomain had confirmed sequence reported in GenBank. The fusion protein had the immunoreactivity. Conclusion Human thyroid stimulating hormone receptor was successfully cloned in eukaryotic expression vector and the co