由于腺病毒载体转导目的基因高效率、低致病性、高滴度以及在体内不整合入宿主细胞染色体等优点,腺病毒载体已被认为是最为有效的转基因载体之一,并广泛运用于人类基因治疗。但是腺病毒载体还存在许多不足,故在第一、第二代腺病毒载体...由于腺病毒载体转导目的基因高效率、低致病性、高滴度以及在体内不整合入宿主细胞染色体等优点,腺病毒载体已被认为是最为有效的转基因载体之一,并广泛运用于人类基因治疗。但是腺病毒载体还存在许多不足,故在第一、第二代腺病毒载体的基础上发展了第三代腺病毒载体,通常称为辅助病毒依赖型腺病毒或无肠腺病毒或具有高包装能力的腺病毒(high-capacity or gutted orhelp-dependent Adenovirus,HD-AdV),现将第三代腺病毒载体的特性研究进展作一综述。展开更多
The Cre-loxP system was introduced in theproduction of recombinant AAV (rAAV). Defective adenovi-rus AdLC was used as the helper virus. While doing themass production of recombinant AAV carrying EGFP orhuman clotting ...The Cre-loxP system was introduced in theproduction of recombinant AAV (rAAV). Defective adenovi-rus AdLC was used as the helper virus. While doing themass production of recombinant AAV carrying EGFP orhuman clotting factor Ⅸ (hFⅨ) gene, the generation ofhelper virus was significantly limited, it increased the sim-plicity of rAAV purification. The results from in vivo study demonstrated the superiority of this method. This systemprovides a novel approach for the application of rAAV sys-tem in gene therapy.展开更多
The recombinant adeno-associated viral vector with human coagulation Factor Ⅸ minigene which wasregulated by CMV promoter was constructed. Largequantity of recombinant adeno-associated viral particles (rAAV/ hFⅨ) wa...The recombinant adeno-associated viral vector with human coagulation Factor Ⅸ minigene which wasregulated by CMV promoter was constructed. Largequantity of recombinant adeno-associated viral particles (rAAV/ hFⅨ) was prepared by the HSV/AAV hybrid helper virus method. Southern dot blot assay and QC-PCR indicated that the titer of the virus was 3.6×1012 v.g./mL. It demonstrated that this method can effectively overcome the hurdles of mass production of AAV vector. Followed by anintramuscular injection of viral vectors (7.5×1011 v.g./mouse) in the quadriceps femoris, an elevation of human Factor Ⅸexpression in the plasma of hemophilia B mice was detected (387 ng/mL) and persisted more than 12 weeks. The level of anti-virus antibody in plasma aligned with the Factor Ⅸexpression curve. The QC-PCR method is easier and moreaccurate than traditional dot hybridization fordetermination of the titer of recombinant adeno-associated virus. Moreover, there are no HSV particles existing inproduced AAV assayed by RT-PCR. AAV is the only virus that has been amplified from AAV-injected muscle by PCR.展开更多
文摘由于腺病毒载体转导目的基因高效率、低致病性、高滴度以及在体内不整合入宿主细胞染色体等优点,腺病毒载体已被认为是最为有效的转基因载体之一,并广泛运用于人类基因治疗。但是腺病毒载体还存在许多不足,故在第一、第二代腺病毒载体的基础上发展了第三代腺病毒载体,通常称为辅助病毒依赖型腺病毒或无肠腺病毒或具有高包装能力的腺病毒(high-capacity or gutted orhelp-dependent Adenovirus,HD-AdV),现将第三代腺病毒载体的特性研究进展作一综述。
基金the National 63High-tech Development Program (Grant No. 2001AA217181)the Na-tional Natural Science Foundation of China (Grant No. 39880019)Foundation of Doctor Degree Thesis from the Ministry of Education of China (Grant No. 199925) and Doctor Deg
文摘The Cre-loxP system was introduced in theproduction of recombinant AAV (rAAV). Defective adenovi-rus AdLC was used as the helper virus. While doing themass production of recombinant AAV carrying EGFP orhuman clotting factor Ⅸ (hFⅨ) gene, the generation ofhelper virus was significantly limited, it increased the sim-plicity of rAAV purification. The results from in vivo study demonstrated the superiority of this method. This systemprovides a novel approach for the application of rAAV sys-tem in gene therapy.
基金supported by the National Natural Science Foundation of China(Grant No.30100102).
文摘The recombinant adeno-associated viral vector with human coagulation Factor Ⅸ minigene which wasregulated by CMV promoter was constructed. Largequantity of recombinant adeno-associated viral particles (rAAV/ hFⅨ) was prepared by the HSV/AAV hybrid helper virus method. Southern dot blot assay and QC-PCR indicated that the titer of the virus was 3.6×1012 v.g./mL. It demonstrated that this method can effectively overcome the hurdles of mass production of AAV vector. Followed by anintramuscular injection of viral vectors (7.5×1011 v.g./mouse) in the quadriceps femoris, an elevation of human Factor Ⅸexpression in the plasma of hemophilia B mice was detected (387 ng/mL) and persisted more than 12 weeks. The level of anti-virus antibody in plasma aligned with the Factor Ⅸexpression curve. The QC-PCR method is easier and moreaccurate than traditional dot hybridization fordetermination of the titer of recombinant adeno-associated virus. Moreover, there are no HSV particles existing inproduced AAV assayed by RT-PCR. AAV is the only virus that has been amplified from AAV-injected muscle by PCR.
