[Expression of SARS coronavirus nucleocapsid protein and construction of its DNA vaccine].

Aim: To express the nucleocapsid (N) protein of SARS coronavirus (SARS-CoV) in E. coli and construct its DNA vaccine.

Methods: The prokaryotic expression vector pQEN containing N gene was constructed and transformed into the E.

[Prokaryotic expression of SARS coronavirus spike protein and construction of its DNA vaccine].

Objective: To study the immunological characteristics of the spike (S) protein of SARS coronavirus (SARS-CoV) and analyze the feasibility of using this protein as the component for SARS vaccine development.

Methods: The two truncated fragments of S gene were separately cloned into the prokaryotic expression vector pET-15b and expressed in E.coli.

[Expression, refolding and biological activity of recombinant type-I metalloproteinase acutolysin a from Agkistrodon acutus].

Type-I snake venom metalloproteinase acutolysin A gene was cloned into the prokaryotic expression vector pBAD/gIIIA and the resulting recombinant plasmid pDS was obtained. By the induction with 0.02% L-(+)-arabinose, the recombinant metalloproteinase was expressed in insoluble inclusion body in E.

[Construction and in vitro activity of specific dual-ribozyme against alpha 1 (I) and (III) procollagen genes].

Overexpression of procollagen gene can cause the extraordinary increase of collagen's synthesis and therefore lead to the keloid and hypertrophic scar. To utilize ribozyme to suppress the expression of procollagen genes, a eukaryotic expression recombinant plasmid containing a dual-ribozyme gene against alpha 1 (I) and alpha 1 (III) procollagen genes was constructed. The ribozyme from in vitro transcription was incubated with target transcripts from recombinant plasmids which separately contained the fragments of the second exons of pro alpha 1 (I) and pro alpha 1 (III) collagen genes under various experimental conditions.

Construction of HBV-specific ribozyme and its recombinant with HDV and their cleavage activity in vitro.

AIM:To construct the recombinant of HDV cDNA and HBV specific ribozyme gene by recombinant PCR in order to use HDV as a transporting vector carrying HBV-specific ribozyme into liver cells for inhibiting the replication of HBV.METHODS:We separately cloned the ribozyme (RZ) gene and recombinant DVRZ (comprising HDV cDNA and HBV-specific ribozyme gene) into the downstream of T7 promoter of pTAdv-T vector and studied the in vitro cleavage activity of their transcripts (rRZ, rDVRZ) on target RNA (rBVCF) from in vitro transcription of HBV C gene fragment(BVCF).RESULTS:Both the simple (rRZ) and the recombinant ribozyme rDVRZ could efficiently catalyze the cleavage of target RNA (rBVCF) under different temperatures (37°, 42° and 55°) and Mg(2+) concentrations (10mmol/L, 15mmol/L and 20mmol/L) and their catalytic activity tended to increase as the temperature was rising.