PDF(Sci-hub)
Abstract：
A major obstacle to creating precisely expressed transgenes lies in the epigenetic effects of the host chromatin that surrounds them. Here we present a strategy to overcome this problem, employing a Gal4-inducible luciferase assay to systematically quantify position effects of host chromatin and the ability of insulators to counteract these effects at phiC31 integration loci randomly distributed throughout the Drosophila genome. We identify loci that can be exploited to deliver precise doses of transgene expression to specific tissues. Moreover, we uncover a previously unrecognized property of the gypsy retrovirus insulator to boost gene expression to levels severalfold greater than at most or possibly all un-insulated loci, in every tissue tested. These findings provide the first opportunity to create a battery of transgenes that can be reliably expressed at high levels in virtually any tissue by integration at a single locus, and conversely, to engineer a controlled phenotypic allelic series by exploiting several loci. The generality of our approach makes it adaptable to other model systems to identify and modify loci for optimal transgene expression.
摘要：
创建精确表达的转基因的主要障碍在于围绕它们的宿主染色质的表观遗传效应。在这里，我们提出了一个克服这个问题的策略，采用Gal4诱导型荧光素酶测定法来系统地定量宿主染色质的位置效应以及绝缘子在整个果蝇基因组中随机分布的phiC31整合基因座上抵消这些效应的能力。我们鉴定了可用于向特定组织递送精确剂量的转基因表达的基因座。此外，我们发现了吉普赛逆转录病毒绝缘子的一个以前未被认识到的特性，以将基因表达提高到比最多或可能所有未绝缘基因座高几倍的水平，在每个测试的组织中。这些发现为创造一系列转基因提供了第一个机会，这些转基因可以通过整合在单个基因座在几乎任何组织中以高水平可靠表达。反过来,通过利用几个基因座来设计受控的表型等位基因系列。我们方法的一般性使其适用于其他模型系统以鉴定和修饰基因座以实现最佳转基因表达。
