Various applications of TALEN- and CRISPR/Cas9-mediated homologous recombination to modify the Drosophila genome.
Various applications of TALEN- and CRISPR/Cas9-mediated homologous recombination to modify the Drosophila genome.
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TALEN 和 CRISPR/Cas9 介导的同源重组在修改果蝇基因组方面的各种应用
DOI：
10.1242/bio.20147682
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发表时间：
2014-04-15
期刊：
影响因子：
2.4
通讯作者：
Jiao R
中科院分区：
文献类型：
--
作者：
Yu Z;Chen H;Liu J;Zhang H;Yan Y;Zhu N;Guo Y;Yang B;Chang Y;Dai F;Liang X;Chen Y;Shen Y;Deng WM;Chen J;Zhang B;Li C;Jiao R
修改许多生物体的基因组变得像在试管中操纵DNA一样容易，这是通过两种最近开发的技术实现的，这两种技术基于可定制的DNA结合蛋白TALEN或CRISPR/Cas9系统。在这里，我们描述了一系列有效的应用程序来自这两种技术，结合各种同源供体DNA质粒，操纵果蝇基因组：（1）精确产生基因组缺失;（2）在单核苷酸分辨率的DNA片段的基因组替换;（3）产生精确的插入标记靶蛋白，用于跟踪其内源性表达。为了更方便的基因组操作，我们通过同源重组敲入白色标记物建立了一个易于筛选的平台。此外，我们提供了一种策略，以消除在“末端”重组过程中产生的不需要的重复。我们的研究结果还表明，TALEN和CRISPR/Cas9在通过HDR（同源定向修复）介导基因组修饰方面具有相当的效率; TALEN或CRISPR/Cas9系统都可以有效地介导果蝇体内高达5 kb的DNA片段的替换，为果蝇基因组的功能注释提供了理想的遗传工具。
ABSTRACT Modifying the genomes of many organisms is becoming as easy as manipulating DNA in test tubes, which is made possible by two recently developed techniques based on either the customizable DNA binding protein, TALEN, or the CRISPR/Cas9 system. Here, we describe a series of efficient applications derived from these two technologies, in combination with various homologous donor DNA plasmids, to manipulate the Drosophila genome: (1) to precisely generate genomic deletions; (2) to make genomic replacement of a DNA fragment at single nucleotide resolution; and (3) to generate precise insertions to tag target proteins for tracing their endogenous expressions. For more convenient genomic manipulations, we established an easy-to-screen platform by knocking in a white marker through homologous recombination. Further, we provided a strategy to remove the unwanted duplications generated during the “ends-in” recombination process. Our results also indicate that TALEN and CRISPR/Cas9 had comparable efficiency in mediating genomic modifications through HDR (homology-directed repair); either TALEN or the CRISPR/Cas9 system could efficiently mediate in vivo replacement of DNA fragments of up to 5 kb in Drosophila, providing an ideal genetic tool for functional annotations of the Drosophila genome.
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影响因子：
7
作者：
Auer TO;Duroure K;De Cian A;Concordet JP;Del Bene F
通讯作者：
Del Bene F
影响因子：
7
作者：
Piganeau M;Ghezraoui H;De Cian A;Guittat L;Tomishima M;Perrouault L;René O;Katibah GE;Zhang L;Holmes MC;Doyon Y;Concordet JP;Giovannangeli C;Jasin M;Brunet E
通讯作者：
Brunet E
影响因子：
56.9
作者：
Jinek, Martin;Chylinski, Krzysztof;Charpentier, Emmanuelle
通讯作者：
Charpentier, Emmanuelle
影响因子：
7
作者：
Maresca M;Lin VG;Guo N;Yang Y
通讯作者：
Yang Y
通过简单的 TALEN 策略对果蝇基因组进行高效、特异性的修饰
影响因子：
5.9
作者：
Liu, Jiyong;Li, Changqing;Jiao, Renjie
通讯作者：
Jiao, Renjie