Zinc finger recombinases with adaptable DNA sequence specificity.
Zinc finger recombinases with adaptable DNA sequence specificity.
复制标题
DOI：
10.1371/journal.pone.0019537
复制
发表时间：
2011-04-29
期刊：
影响因子：
3.7
通讯作者：
Stark WM
中科院分区：
文献类型：
--
作者：
Proudfoot C;McPherson AL;Kolb AF;Stark WM
位点特异性重组酶已经成为遗传学和分子生物学中用于精确切除或整合DNA序列的重要工具。然而，它们的效用目前仅限于重组酶识别的位点已被引入被操纵的DNA中或天然“假位点”已经存在的情况。如果重组酶的活性可以靶向天然基因组DNA中的选定序列，许多新的应用将变得可行。在这里，我们证明了有效的位点特异性重组的几个序列取自1.9个酶对基因座的生物技术的兴趣（在牛β-酪蛋白基因），介导的锌指重组酶（ZFR），嵌合酶与连接的锌指（DNA识别）和重组酶（催化）结构域。在这里测试的“Z-位点”中，22 bp酪蛋白基因序列侧接由锌指结构域识别的9 bp基序。不对称的Z-网站重组的同时行动的两个ZFR具有不同的锌指DNA结合特异性，并可以重组与异源位点的存在下，第三重组酶。我们的研究结果表明，工程化的ZFR可以被设计为促进在许多天然DNA序列的位点特异性重组。
Site-specific recombinases have become essential tools in genetics and molecular biology for the precise excision or integration of DNA sequences. However, their utility is currently limited to circumstances where the sites recognized by the recombinase enzyme have been introduced into the DNA being manipulated, or natural ‘pseudosites’ are already present. Many new applications would become feasible if recombinase activity could be targeted to chosen sequences in natural genomic DNA. Here we demonstrate efficient site-specific recombination at several sequences taken from a 1.9 kilobasepair locus of biotechnological interest (in the bovine β-casein gene), mediated by zinc finger recombinases (ZFRs), chimaeric enzymes with linked zinc finger (DNA recognition) and recombinase (catalytic) domains. In the "Z-sites" tested here, 22 bp casein gene sequences are flanked by 9 bp motifs recognized by zinc finger domains. Asymmetric Z-sites were recombined by the concomitant action of two ZFRs with different zinc finger DNA-binding specificities, and could be recombined with a heterologous site in the presence of a third recombinase. Our results show that engineered ZFRs may be designed to promote site-specific recombination at many natural DNA sequences.
登录
查看更多内容
影响因子：
46.9
作者：
Isalan, M;Klug, A;Choo, Y
通讯作者：
Choo, Y
影响因子：
14.9
作者：
Olorunniji FJ;He J;Wenwieser SV;Boocock MR;Stark WM
通讯作者：
Stark WM
DOI：
10.1073/pnas.1533177100
发表时间：
2003-07-22
影响因子：
11.1
作者：
Akopian, A;He, JY;Stark, WM
通讯作者：
Stark, WM
影响因子：
5.6
作者：
Zaccolo, M;Williams, DM;Gherardi, E
通讯作者：
Gherardi, E
影响因子：
3.2
作者：
CHANG, ACY;COHEN, SN
通讯作者：
COHEN, SN