Effect of Genome Size on AAV Vector Packaging
Effect of Genome Size on AAV Vector Packaging
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DOI：
10.1038/mt.2009.255
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发表时间：
2010-01-01
影响因子：
12.4
通讯作者：
Colosi, Peter
中科院分区：
文献类型：
--
作者：
Wu, Zhijian;Yang, Hongyan;Colosi, Peter
腺相关病毒（AAV）载体基因组的长度被限制在5千碱基（kb），因为它们的包装极限被认为与亲本AAV基因组的大小相似。最近的报告声称，更大的载体基因组可以完整地包装。我们检测了来自质粒编码的AAV载体的包装载体基因组，长度从4.7到8.7 kb不等，使用AAV类型2、5和8衣壳。Southern blot分析表明，无论质粒编码载体大小或衣壳类型如何，包装后的AAV载体基因组长度均未超过5.2 kb。这一结果被用链特异性寡核苷酸的载体基因组探测证实。由超过5kb的质粒编码载体衍生的包装载体基因组在长度上是异质的，并且在5'端被截断。尽管它们的基因组被截断，但质粒编码载体制备的载体在体外高感染多重性（MOI）下介导了超过5.2 kb的报告基因表达。其表达效率明显低于带基因组的报告载体
Adeno-associated virus (AAV) vector genomes have been limited to 5 kilobases (kb) in length because their packaging limit was thought to be similar to the size of the parent AAV genome. Recent reports claim that significantly larger vector genomes can be packaged intact. We examined the packaged vector genomes from plasmid-encoded AAV vectors that ranged from 4.7 to 8.7 kb in length, using AAV types 2, 5, and 8 capsids. Southern blot analysis indicated that packaged AAV vector genomes never exceeded 5.2 kb in length irrespective of the size of the plasmid-encoded vector or the capsid type. This result was confirmed by vector genome probing with strand-specific oligonucleotides. The packaged vector genomes derived from plasmid-encoded vectors exceeding 5 kb were heterogeneous in length and truncated on the 5' end. Despite their truncated genomes, vector preparations produced from plasmid-encoded vectors exceeding 5.2 kb mediated reporter gene expression in vitro at high multiplicity of infection (MOI). The efficiency of expression was substantially lower than that of reporter vectors with genomes