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. But the activity of rRZ was evidently higher than that of rDVRZ.CONCLUSION:The recombinant of HDV cDNA and ribozyme gene had the potential of being further explored and used in gene therapy of HBV infection.
Wen
Shu Juan
SJ
Xiang
Kai-Jun
KJ
Huang
Zhen-Hua
ZH
Zhou
Rong
R
Qi
Xue-Zhong
XZ