Scarless engineering of the Drosophila genome near any site-specific integration site
Scarless engineering of the Drosophila genome near any site-specific integration site
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DOI：
10.1093/genetics/iyab012
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发表时间：
2021-01-23
期刊：
影响因子：
3.3
通讯作者：
Mann, Richard S.
中科院分区：
文献类型：
--
作者：
Feng, Siqian;Lu, Shan;Mann, Richard S.
我们描述了一种简单而有效的技术，它允许在任何含有倒置attP盒的着陆点附近对果蝇基因组序列进行无疤痕工程，例如模拟插入。这种两步法结合了phiC31整合酶介导的位点特异性整合和归巢核酸酶介导的局部复制的解析，有效地将原始着陆位点的等位基因转换为只有期望变化的修饰等位基因(S)。这种方法结合了显性标记，使得在每一步都能有效地识别正确的个体苍蝇。原则上，单一的ATP站点和FRT站点也是有效的着陆点。鉴于苍蝇群落中可用的着陆点数量越来越多，这种方法提供了一种简单而快速的方法，以无疤痕的方式有效地编辑大多数果蝇基因组。这项技术也应该适用于其他物种。
We describe a simple and efficient technique that allows scarless engineering of Drosophila genomic sequences near any landing site containing an inverted attP cassette, such as a MiMIC insertion. This two-step method combines phiC31 integrase-mediated site-specific integration and homing nuclease-mediated resolution of local duplications, efficiently converting the original landing site allele to modified alleles that only have the desired change(s). Dominant markers incorporated into this method allow correct individual flies to be efficiently identified at each step. In principle, single attP sites and FRT sites are also valid landing sites. Given the large and increasing number of landing site lines available in the fly community, this method provides an easy and fast way to efficiently edit the majority of the Drosophila genome in a scarless manner. This technique should also be applicable to other species.