Chromosomal deletions and inversions mediated by TALENs and CRISPR/Cas in zebrafish.
Chromosomal deletions and inversions mediated by TALENs and CRISPR/Cas in zebrafish.
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DOI：
10.1093/nar/gkt464
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发表时间：
2013-08
影响因子：
14.9
通讯作者：
Zhang B
中科院分区：
文献类型：
--
作者：
Xiao A;Wang Z;Hu Y;Wu Y;Luo Z;Yang Z;Zu Y;Li W;Huang P;Tong X;Zhu Z;Lin S;Zhang B
定制的TALEN和Cas9/gRNA已用于斑马鱼中的靶向诱变，以将indel诱导到蛋白质编码基因中。然而，插入缺失通常不足以破坏非编码基因、基因簇或调控序列的功能，而大的基因组缺失或倒位对于该目的更理想。通过将两对TALEN mRNA或两种gRNA与靶向相同染色体的远端DNA位点的Cas9 mRNA一起注射，我们获得了大小范围从几百个碱基到近1 Mb的可预测的基因组缺失或倒位。我们已经成功地通过TALEN对11个染色体基因座进行了这种类型的修饰，并通过Cas9/gRNA对不同的gRNA对组合（包括miRNA和蛋白质编码基因簇）对2个染色体基因座进行了这种类型的修饰。八个TALEN靶向株系中有七个传递了缺失，一个通过种系传递了倒位。我们的研究结果表明，TALEN和Cas9/gRNA都可以作为一种有效的工具来工程化基因组，以实现大的缺失或倒位，包括覆盖多个基因和非编码序列的片段。为了促进现有ZFN、TALEN和CRISPR/Cas数据的分析和应用，我们更新了EENdb数据库，以提供所有报告的靶向人类和斑马鱼基因组的工程核酸内切酶的染色体视图。
Customized TALENs and Cas9/gRNAs have been used for targeted mutagenesis in zebrafish to induce indels into protein-coding genes. However, indels are usually not sufficient to disrupt the function of non-coding genes, gene clusters or regulatory sequences, whereas large genomic deletions or inversions are more desirable for this purpose. By injecting two pairs of TALEN mRNAs or two gRNAs together with Cas9 mRNA targeting distal DNA sites of the same chromosome, we obtained predictable genomic deletions or inversions with sizes ranging from several hundred bases to nearly 1 Mb. We have successfully achieved this type of modifications for 11 chromosomal loci by TALENs and 2 by Cas9/gRNAs with different combinations of gRNA pairs, including clusters of miRNA and protein-coding genes. Seven of eight TALEN-targeted lines transmitted the deletions and one transmitted the inversion through germ line. Our findings indicate that both TALENs and Cas9/gRNAs can be used as an efficient tool to engineer genomes to achieve large deletions or inversions, including fragments covering multiple genes and non-coding sequences. To facilitate the analyses and application of existing ZFN, TALEN and CRISPR/Cas data, we have updated our EENdb database to provide a chromosomal view of all reported engineered endonucleases targeting human and zebrafish genomes.
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影响因子：
56.9
作者：
Jinek, Martin;Chylinski, Krzysztof;Charpentier, Emmanuelle
通讯作者：
Charpentier, Emmanuelle
影响因子：
14.9
作者：
Flicek P;Ahmed I;Amode MR;Barrell D;Beal K;Brent S;Carvalho-Silva D;Clapham P;Coates G;Fairley S;Fitzgerald S;Gil L;García-Girón C;Gordon L;Hourlier T;Hunt S;Juettemann T;Kähäri AK;Keenan S;Komorowska M;Kulesha E;Longden I;Maurel T;McLaren WM;Muffato M;Nag R;Overduin B;Pignatelli M;Pritchard B;Pritchard E;Riat HS;Ritchie GR;Ruffier M;Schuster M;Sheppard D;Sobral D;Taylor K;Thormann A;Trevanion S;White S;Wilder SP;Aken BL;Birney E;Cunningham F;Dunham I;Harrow J;Herrero J;Hubbard TJ;Johnson N;Kinsella R;Parker A;Spudich G;Yates A;Zadissa A;Searle SM
通讯作者：
Searle SM
影响因子：
14.9
作者：
Cade L;Reyon D;Hwang WY;Tsai SQ;Patel S;Khayter C;Joung JK;Sander JD;Peterson RT;Yeh JR
通讯作者：
Yeh JR
影响因子：
46.9
作者：
Cho, Seung Woo;Kim, Sojung;Kim, Jin-Soo
通讯作者：
Kim, Jin-Soo
影响因子：
46.9
作者：
Kim, Yongsub;Kweon, Jiyeon;Kim, Jin-Soo
通讯作者：
Kim, Jin-Soo