Construction of a novel human artificial chromosome vector for gene delivery
Construction of a novel human artificial chromosome vector for gene delivery
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DOI：
10.1016/j.bbrc.2004.06.145
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发表时间：
2004-08-20
影响因子：
3.1
通讯作者：
Oshimura, M
中科院分区：
文献类型：
--
作者：
Katoh, M;Ayabe, F;Oshimura, M
常规转基因的潜在问题包括宿主基因组的插入破坏和通过随机整合的转基因的不可预测的、不可再现的表达。或者，人类人工染色体（HACs）可以避免一些问题。虽然产生了几种HAC，并评估了它们的有丝分裂稳定性，但通过HAC引入外源基因的实用方法还有待探索。在这项研究中，我们开发了一种新的HAC从序列就绪的人类21号染色体端粒定向染色体截短，并添加了一个loxP序列的环状DNA的位点特异性插入的Cre/loxP系统。该21 HAC载体通过微细胞融合递送至人细胞系HT 1080，结合着丝粒蛋白A、B和C，并且在长期培养期间在无选择的情况下是有丝分裂稳定的。插入HAC载体的EGFP基因持续表达。这些结果表明，HAC载体提供了一个有用的系统，在同基因细胞系的基因功能的研究。(C)2004爱思唯尔公司All rights reserved.
Potential problems of conventional transgenes include insertional disruption of the host genome and unpredictable, irreproducible expression of the transgene by random integration. Alternatively, human artificial chromosomes (HACs) can circumvent some of the problems. Although several HACs were generated and their mitotic stability was assessed, a practical way for introducing exogenous genes by the HACs has yet to be explored. In this study, we developed a novel HAC from sequence-ready human chromosome 21 by telomere-directed chromosome truncation and added a loxP sequence for site-specific insertion of circular DNA by the Cre/loxP system. This 21 HAC vector, delivered to a human cell line HT1080 by microcell fusion, bound centromere proteins A, B, and C and was mitotically stable during long-term culture without selection. The EGFP gene inserted in the HAC vector expressed persistently. These results suggest that the HAC vector provides useful system for functional studies of genes in isogenic cell lines. (C) 2004 Elsevier Inc. All rights reserved.