Transcription Activator-like Effector Nuclease

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Scheme of using TALEN

A transcription activator-like effector nuclease (dt. ' Transcription activator-like effector nuclease ', TALEN) is an artificial sequence-specific restriction enzyme .

properties

Transcription activator-like effector nucleases are fusion proteins from a TAL effector - DNA-binding domain and an endonuclease domain . The sequence-specific binding occurs through the DNA-binding domain usedin the course of a protein design, while the endonuclease results in a comparatively little sequence-specific cut. In addition to the CRISPR / Cas method and zinc finger nucleases, TALENs aremethods of genome editing .

The valley effector domain originates from Xanthomonas bacteria and has a conserved sequence of 33 to 34 amino acids. Positions 12 and 13, on the other hand, are variable ( repeat variable direction ) and are responsible for the recognition of nucleotides in DNA. A lack of specificity of the Tal effector domain leads to double-strand breaks in other DNA sequences.

For use in yeast, plant or animal cells, the endonuclease domain from FokI is mostly used. While the wild type of FokI was originally used, various mutations have subsequently been introduced to improve the specificity and activity in FokI . FokI is a dimeric protein and the number of base pairs between the valley effector domain and the two FokI subunits affects its activity.

The DNA sequence of the designed TALEN fusion protein is cloned into a plasmid and then transfected into the target cells . The gene is translated into the protein in the cell and transported into the cell nucleus by the signal sequence it contains , where it unfolds its activity.

Web links

Individual evidence

  1. ↑ Jens Boch: TALEs of genome targeting . In: Nature Biotechnology . 29, No. 2, February 2011, pp. 135-6. doi : 10.1038 / nbt.1767 . PMID 21301438 .
  2. ^ J. Boch, H. Scholze, S. Schornack, A. Landgraf, S. Hahn, S. Kay, T. Lahaye, A. Nickstadt, U. Bonas: Breaking the code of DNA binding specificity of TAL-type III effectors . In: Science. Volume 326, number 5959, December 2009, ISSN  1095-9203 , pp. 1509-1512, doi : 10.1126 / science.1178811 . PMID 19933107 .
  3. ↑ MJ Moscou, AJ Bogdanove: A simple cipher governs DNA recognition by TAL effectors. In: Science. Volume 326, number 5959, December 2009, ISSN  1095-9203 , p. 1501, doi : 10.1126 / science.1178817 . PMID 19933106 .
  4. ^ A b C. Mussolino, R. Morbitzer, F. Lütge, N. Dannemann, T. Lahaye, T. Cathomen: A novel TALE nuclease scaffold enables high genome editing activity in combination with low toxicity. In: Nucleic acids research. Volume 39, Number 21, November 2011, pp. 9283-9293, ISSN  1362-4962 . doi : 10.1093 / nar / gkr597 . PMID 21813459 . PMC 3241638 (free full text).
  5. ↑ M. Christian, T. Cermak, EL Doyle, C. Schmidt, F. Zhang, A. Hummel, AJ Bogdanove, DF Voytas: Targeting DNA double-strand breaks with TAL effector nucleases. In: Genetics. Volume 186, Number 2, October 2010, ISSN  1943-2631 , pp. 757-761, doi : 10.1534 / genetics.110.120717 . PMID 20660643 , PMC 2942870 (free full text).
  6. ↑ T. Li, S. Huang, WZ Jiang, D. Wright, MH Spalding, DP Weeks, B. Yang: TAL nucleases (TALNs): hybrid proteins composed of TAL effectors and FokI DNA-cleavage domain. In: Nucleic acids research. Volume 39, Number 1, January 2011, ISSN  1362-4962 , pp. 359-372, doi : 10.1093 / nar / gkq704 . PMID 20699274 , PMC 3017587 (free full text).
  7. ↑ MM Mahfouz, L. Li, M. Shamimuzzaman, A. Wibowo, X. Fang, JK Zhu: De novo-engineered transcription activator-like effector (TALE) hybrid nuclease with novel DNA binding specificity creates double-strand breaks. In: Proceedings of the National Academy of Sciences . Volume 108, Number 6, February 2011, ISSN  1091-6490 , pp. 2623-2628, doi : 10.1073 / pnas.1019533108 . PMID 21262818 , PMC 3038751 (free full text).
  8. ↑ a b T. Cermak, EL Doyle, M. Christian, L. Wang, Y. Zhang, C. Schmidt, JA Baller, NV Somia, AJ Bogdanove, DF Voytas: Efficient design and assembly of custom TALEN and other TAL effector- based constructs for DNA targeting. In: Nucleic acids research. Volume 39, number 12, July 2011, p. E82, ISSN  1362-4962 . doi : 10.1093 / nar / gkr218 . PMID 21493687 . PMC 3130291 (free full text).
  9. ↑ a b JC Miller, S. Tan, G. Qiao, KA Barlow, J. Wang, DF Xia, X. Meng, DE Paschon, E. Leung, SJ Hinkley, GP Dulay, KL Hua, I. Ankoudinova, GJ Cost , FD Urnov, HS Zhang, MC Holmes, L. Zhang, PD Gregory, EJ Rebar: A TALE nuclease architecture for efficient genome editing. In: Nature Biotechnology . Volume 29, Number 2, February 2011, ISSN  1546-1696 , pp. 143-148, doi : 10.1038 / nbt.1755 . PMID 21179091 .
  10. ↑ D. Hockemeyer, H. Wang, S. Kiani, CS Lai, Q. Gao, JP Cassady, GJ Cost, L. Zhang, Y. Santiago, JC Miller, B. Zeitler, JM Cherone, X. Meng, SJ Hinkley , EJ Rebar, PD Gregory, FD Urnov, R. Jaenisch: Genetic engineering of human pluripotent cells using TALE nucleases. In: Nature Biotechnology . Volume 29, Number 8, August 2011, pp. 731-734, ISSN  1546-1696 . doi : 10.1038 / nbt.1927 . PMID 21738127 . PMC 3152587 (free full text).
  11. ↑ AJ Wood, TW Lo, B. Zeitler, CS Pickle, EJ Ralston, AH Lee, R. Amora, JC Miller, E. Leung, X. Meng, L. Zhang, EJ Rebar, PD Gregory, FD Urnov, BJ Meyer : Targeted genome editing across species using ZFNs and TALENs. In: Science. Volume 333, Number 6040, July 2011, p. 307, ISSN  1095-9203 . doi : 10.1126 / science.1207773 . PMID 21700836 . PMC 3489282 (free full text).
  12. ↑ Y. Doyon, TD Vo, MC Mendel, SG Greenberg, J. Wang, DF Xia, JC Miller, FD Urnov, PD Gregory, MC Holmes: Enhancing zinc-finger-nuclease activity with improved obligate heterodimeric architectures. In: Nature methods. Volume 8, Number 1, January 2011, pp. 74-79, ISSN  1548-7105 . doi : 10.1038 / nmeth.1539 . PMID 21131970 .
  13. ↑ M. Szczepek, V. Brondani, J. Büchel, L. Serrano, DJ Segal, T. Cathomen: Structure-based redesign of the dimerization interface reduces the toxicity of zinc-finger nucleases. In: Nature Biotechnology . Volume 25, Number 7, July 2007, pp. 786-793, ISSN  1087-0156 . doi : 10.1038 / nbt1317 . PMID 17603476 .
  14. ↑ J. Guo, T. Gaj, CF Barbas: Directed evolution of an enhanced and highly efficient FokI cleavage domain for zinc finger nucleases. In: Journal of molecular biology. Volume 400, Number 1, July 2010, pp. 96-107, ISSN  1089-8638 . doi : 10.1016 / j.jmb.2010.04.060 . PMID 20447404 . PMC 2885538 (free full text).