Self-complementary recombinant adeno-associated virus (scAAV) vectors promote efficient transduction independently of DNA synthesis
Self-complementary recombinant adeno-associated virus (scAAV) vectors promote efficient transduction independently of DNA synthesis
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DOI：
10.1038/sj.gt.3301514
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发表时间：
2001-08-01
期刊：
影响因子：
5.1
通讯作者：
Samulski, RJ
中科院分区：
文献类型：
--
作者：
McCarty, DM;Monahan, PE;Samulski, RJ
腺相关病毒（AAV）载体包装单链基因组，需要宿主细胞合成互补链进行转导。然而，当基因组是野生型大小的一半时，AAV可以包装两个拷贝或二聚体反向重复DNA分子。二聚体或自身互补分子（scAAV）应该自发地重新退火，减轻宿主细胞DNA合成的需要。我们生成并表征了scAAV载体，以绕过第二链合成的限速步骤。在体外，scAAV载体是比常规rAAV更有效的5至140倍的转导剂，具有5.9：1的颗粒与转导单位比。这种效率既没有大大增加与广告的共感染，也没有受到抑制的羟基脲，表明转导是独立的DNA合成。在体内，表达促红细胞生成素的scAAV导致比常规单链载体更快和更高水平的红细胞比容。这些新的scAAV载体代表了rAAV转导中的生化中间体，并且应该为载体转导的生物学提供新的见解。
Adeno-associated virus (AAV) vectors package single-stranded genomes and require host-cell synthesis of the complementary strand for transduction. However, when the genome is half wild-type size, AAV can package either two copies, or dimeric inverted repeat DNA molecules. Dimeric, or self-complementary molecules (scAAV) should spontaneously reanneal, alleviating the requirement for host-cell DNA synthesis. We generated and characterized scAAV vectors in order to bypass the rate-limiting step of second-strand synthesis. In vitro, scAAV vectors were five- to 140-fold more efficient transducing agents than conventional rAAV, with a 5.9:1 particle to transducing unit ratio. This efficiency is neither greatly increased by co-infection with Ad, nor inhibited by hydroxyurea, demonstrating that transduction is independent of DNA synthesis. In vivo, scAAV expressing erythropoietin resulted in rapid and higher levels of hematocrit than a conventional single-stranded vector. These novel scAAV vectors represent a biochemical intermediate in rAAV transduction and should provide new insights into the biology of vector transduction.