[Liu, Yun
; Zhao, Hui
; Cheng, Christopher H. K.] Chinese Univ Hong Kong, Sch Biomed Sci, Hong Kong, Hong Kong, Peoples R China
; [Luo, Daji
; Zhu, Zuoyan
; Hu, Wei] Chinese Acad Sci, Inst Hydrobiol, State Key Lab Freshwater Ecol & Biotechnol, Wuhan, Peoples R China
; [Luo, Daji] Wuhan Univ, Sch Basic Med Sci, Dept Genet, Wuhan 430072, Peoples R China
; [Liu, Yun
; Zhao, Hui
; Cheng, Christopher H. K.] Chinese Univ Hong Kong, Shenzhen Res Inst, Sch Biomed Sci, Core Lab, Shenzhen, Peoples R China
Transcription activator-like effector nucleases (TALENs) have so far been applied to disrupt protein-coding genes which constitute only 2-3% of the genome in animals. The majority (70-90%) of the animal genome is actually transcribed as non-coding RNAs (ncRNAs), yet the lack of efficient tools to knockout ncRNA genes hinders studies on their in vivo functions. Here we have developed novel strategies using TALENs to achieve precise and inheritable large genomic deletions and knockout of ncRNA genes in zebrafish. We have demonstrated that individual miRNA genes could be disrupted using one pair of TALENs, whereas large microRNA (miRNA) gene clusters and long non-coding RNA (lncRNA) genes could be precisely deleted using two pairs of TALENs. We have generated large genomic deletions of two miRNA clusters (the 1.2 kb miR-17-92 cluster and the 79.8 kb miR-430 cluster) and one long non-coding RNA (lncRNA) gene (the 9.0 kb malat1), and the deletions are transmitted through the germline. Taken together, our results establish TALENs as a robust tool to engineer large genomic deletions and knockout of ncRNA genes, thus opening up new avenues in the application of TALENs to study the genome in vivo.
英文摘要:
Transcription activator-like effector nucleases (TALENs) have so far been applied to disrupt protein-coding genes which constitute only 2-3% of the genome in animals. The majority (70-90%) of the animal genome is actually transcribed as non-coding RNAs (ncRNAs), yet the lack of efficient tools to knockout ncRNA genes hinders studies on their in vivo functions. Here we have developed novel strategies using TALENs to achieve precise and inheritable large genomic deletions and knockout of ncRNA genes in zebrafish. We have demonstrated that individual miRNA genes could be disrupted using one pair of TALENs, whereas large microRNA (miRNA) gene clusters and long non-coding RNA (lncRNA) genes could be precisely deleted using two pairs of TALENs. We have generated large genomic deletions of two miRNA clusters (the 1.2 kb miR-17-92 cluster and the 79.8 kb miR-430 cluster) and one long non-coding RNA (lncRNA) gene (the 9.0 kb malat1), and the deletions are transmitted through the germline. Taken together, our results establish TALENs as a robust tool to engineer large genomic deletions and knockout of ncRNA genes, thus opening up new avenues in the application of TALENs to study the genome in vivo.
1.Chinese Univ Hong Kong, Sch Biomed Sci, Hong Kong, Hong Kong, Peoples R China 2.Chinese Acad Sci, Inst Hydrobiol, State Key Lab Freshwater Ecol & Biotechnol, Wuhan, Peoples R China 3.Wuhan Univ, Sch Basic Med Sci, Dept Genet, Wuhan 430072, Peoples R China 4.Chinese Univ Hong Kong, Shenzhen Res Inst, Sch Biomed Sci, Core Lab, Shenzhen, Peoples R China
Recommended Citation:
Liu, Yun; Luo, Daji; Zhao, Hui; Zhu, Zuoyan; Hu, Wei; Cheng, Christopher H. K..Inheritable and Precise Large Genomic Deletions of Non-Coding RNA Genes in Zebrafish Using TALENs,PLOS ONE,2013,8(10):e76387
