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AIM To construct the recombinant of HDVcDNA and HBV-specific ribozyme gene byrecombinant PCR in order to use HDV as atransporting vector carrying HBV-specificribozyme into liver cells for inhibiting thereplication of HBV.METHODS We separately cloned the ribozyme(RZ)gene and recombinant DVRZ(comprisingHDV cDNA and HBV-specific ribozyme gene)intothe downstream of T7 promoter of pTAdv-Tvector and studied the in vitro cleavage activityof their transcripts(rRZ,rDVRZ)on target RNA(rBVCF)from in vitro transcription of HBV Cgene fragment(BVCF).RESULTS Both the simple(rRZ)and therecombinant ribozyme rDVRZ could efficientlycatalyze the cleavage of target RNA(rBVCF)under different temperatures(37℃,42℃ and55℃)and Mg~(2+)concentrations(10 mmol/L,15 mmol/L and 20 mmol/L)and their catalyticactivity tended to increase as the temperaturewas rising.But the activity of rRZ was evidentlyhigher than that of rDVRZ.CONCLUSION The recombinant of HDV cDNAand ribozyme gene had the potential of beingfurther explored and used in gene therapy of HBVinfection.
AIM To construct the recombinant of HDV cDNA and HBV-specific ribozyme gene by recombinant PCR in order to use HDV as atransporting vector carrying HBV-specific ribozyme into liver cells for inhibiting thereplication of HBV. METHODS We separately cloned the ribozyme (RZ) gene and recombinant DVRZ ( comprisingHDV cDNA and HBV-specific ribozyme gene) intothe 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 Cgene fragment (BVCF). RESULTS Both the simple (rRZ) and therecombinant ribozyme rDVRZ could efficiently metabolize the cleavage of target RNA (rBVCF) under different temperatures (37 ℃, 42 ℃ and55 ℃) and Mg 2+ concentrations (10 mmol / L, 15 mmol / L and 20 mmol / L) and their catalytic activity tended to increase as the temperature was rising. The activity of rRZ was evidently higher than that of rDVRZ.CONCLUSION The recombinant of HDV cDNA and ribozyme gene had the potential of beingfurthe r explored and used in gene therapy of HBVinfection.