title: [Construction of cTnC-linker-TnI (P) Genes, Expression of Fusion Protein and Preparation of Lyophilized Protein].
abstract: In order to construct and express human cardiac troponin C-linker-troponin I(P) [ cTnC-linker-TnI(P)] fusion protein, detect its activity and prepare lyophilized protein, we searched the CDs of human cTnC and cTnI from GenBank, synthesized cTnC and cTnI(30-110aa) into cloning vector by a short DNA sequence coding for 15 neutral amino acid residues. pCold I-cTnC-linker-TnI(P) was constructed and transformed into E. coli BL21(DE3). Then, cTnC-linker-TnI(P) fusion protein was induced by isopropyl-β-D-thiogalactopyranoside (IPTG). Soluable expression of cTnC-linker-TnI(P) in prokaryotic system was successfully obtained. The fusion protein was purified by Ni²⁺ Sepharose 6 Fast Flow affinity chromatography with over 95% purity and prepared into lyophilized protein. The activity of purified cTnC-linker-TnI(P) and its lyophilized protein were detected by Wondfo Finecare™ cTnI Test. Lyophilized protein of cTnC-linker-TnI(P) was stable for 10 or more days at 37 °C and 4 or more months at 25 °C and 4 °C. The expression system established in this research is feasible and efficient. Lyophilized protein is stable enough to be provided as biological raw materials for further research.
authors.forename: Xiaoli
authors.initials: X
authors.lastname: Song
authors.forename: Xiaoyun
authors.initials: X
authors.lastname: Liu
authors.forename: Lei
authors.initials: L
authors.lastname: Cai
authors.forename: Jianwei
authors.initials: J
authors.lastname: Wu
authors.forename: Jihua
authors.initials: J
authors.lastname: Wang
mesh_terms: D004926:Escherichia coli; D005612:Freeze Drying; D006801:Humans; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis*; D019209:Troponin C / Q000096:biosynthesis*; D019210:Troponin I / Q000096:biosynthesis*
chemical_list: D011993:Recombinant Fusion Proteins; D019209:Troponin C; D019210:Troponin I