Methods 69 (2014) 1
Contents lists available at ScienceDirect
Methods journal homepage: www.elsevier.com/locate/ymeth
Guest Editor’s Introduction
Editing and investigating genomes with TALE and CRISPR/Cas systems: Genome engineering across species using TALENs Genome engineering in cells and organisms to create targeted mutants is a key technology for genetics and biotechnology. The ascent of the mouse as genetic model organism is based on the development of gene targeting technology in embryonic stem (ES) cells that relies on the transfer of vector encoded knockout and knockin mutations into the genome by spontaneous homologous recombination (HR). Besides mouse ES cells, most other cell lines and organisms are refractory to HR so that the available tools enable only random genome integrations. Sequence-specific nucleases for the generation of targeted double-strand breaks revealed as powerful tool for genome engineering, enabling efficient gene editing by the stimulation of DNA repair. The broad application of this technology has long been hampered by the lack of a simple DNA recognition system to construct sequence-specific DNA binding proteins. After decades of research on the structure of DNA binding proteins it seemed unlikely that a simple, modular path of sequence recognition, comprising e.g., four elements each recognizing one deoxyribonucleotide, would exist in nature. Nevertheless, in 2009 a novel, modular DNA recognition code from the transcription activator-like (TAL) proteins of Xanthomonas was discovered and, based on the extensive experience with ZFNs, were rapidly adapted for use as sequence-specific TAL nucleases (TALEN). Within the last two years TAL proteins and TALENs were further characterized and successfully applied for gene editing in various species. To support the use of this new technology by scientists studying the genetics of rice [1,2], mosquitos [3], silk worm [4], cricket [5], fruit fly [6,7], zebrafish [8,9], frog [10], mouse [11,12], rat [13], or of human cells [14] this issue presents step-by-step protocols enabling TALEN-assisted mutagenesis for newcomers. For some species we included multiple independent protocols such that interested users have the choice of selecting a procedure following their individual convenience. The TALEN
http://dx.doi.org/10.1016/j.ymeth.2014.08.012 1046-2023/Ó 2014 Elsevier Inc. All rights reserved.
focused horizon of this issue is further widened by the contents of the companion volume edited by C. Giovannangeli and J.-P. Concordet which presents additional applications of TALEN and TAL DNA binding domains and of its successor technology, the CRISP/Cas system [15]. Altogether, TALENs and CRISP/Cas are at the dawn of a new era of reverse genetics now comprising all species and cell lines derived from them. References [1] [2] [3] [4] [5] [6] [7] [8] [9] [10] [11] [12] [13] [14] [15]
K. Chen, Q. Shan, C. Gao, Methods 69 (2014) 2–8. T. Li, B. Liu, C.Y. Chen, B. Yang, Methods 69 (2014) 9–16. A. Aryan, K.M. Myles, Z.N. Adelman, Methods 69 (2014) 38–45. Y. Takasu, T. Tamura, S. Sajwan, I. Kobayashi, M. Zurovec, Methods 69 (2014) 46–57. T. Watanabe, S. Noji, T. Mito, Methods 69 (2014) 17–21. X. Zhang, I.R.S. Ferreira, F. Schnorrer, Methods 69 (2014) 32–37. J. Liu, Y. Chen, R. Jiao, Methods 69 (2014) 22–31. W.Y. Hwang, R.T. Peterson, J.-R.J. Yeh, Methods 69 (2014) 76–84. P. Huang, A. Xiao, X. Tong, Y. Zu, Z. Wang, B. Zhang, Methods 69 (2014) 67–75. Y. Liu, D. Luo, Y. Lei, W. Hu, H. Zhao, C.H.K. Cheng, Methods 69 (2014) 58–66. Y.H. Sung, Y. Jin, S. Kim, H.-W. Lee, Methods 69 (2014) 85–93. B. Wefers, O. Ortiz, W. Wurst, R. Kühn, Methods 69 (2014) 94–101. S. Ménoret, L. Tesson, S. Rémy, C. Usal, V. Thépenier, R. Thinard, et al, Methods 69 (2014) 102–107. Y.-H. Kim, S. Ramakrishna, H. Kim, J.-S. Kim, Methods 69 (2014) 108–117. C. Giovannangeli, Methods (2014), http://dx.doi.org/10.1016/j.ymeth. 2014.08.013.
Ralf Kühn Benedikt Wefers
Contents lists available at ScienceDirect
Methods journal homepage: www.elsevier.com/locate/ymeth
Guest Editor’s Introduction
Editing and investigating genomes with TALE and CRISPR/Cas systems: Genome engineering across species using TALENs Genome engineering in cells and organisms to create targeted mutants is a key technology for genetics and biotechnology. The ascent of the mouse as genetic model organism is based on the development of gene targeting technology in embryonic stem (ES) cells that relies on the transfer of vector encoded knockout and knockin mutations into the genome by spontaneous homologous recombination (HR). Besides mouse ES cells, most other cell lines and organisms are refractory to HR so that the available tools enable only random genome integrations. Sequence-specific nucleases for the generation of targeted double-strand breaks revealed as powerful tool for genome engineering, enabling efficient gene editing by the stimulation of DNA repair. The broad application of this technology has long been hampered by the lack of a simple DNA recognition system to construct sequence-specific DNA binding proteins. After decades of research on the structure of DNA binding proteins it seemed unlikely that a simple, modular path of sequence recognition, comprising e.g., four elements each recognizing one deoxyribonucleotide, would exist in nature. Nevertheless, in 2009 a novel, modular DNA recognition code from the transcription activator-like (TAL) proteins of Xanthomonas was discovered and, based on the extensive experience with ZFNs, were rapidly adapted for use as sequence-specific TAL nucleases (TALEN). Within the last two years TAL proteins and TALENs were further characterized and successfully applied for gene editing in various species. To support the use of this new technology by scientists studying the genetics of rice [1,2], mosquitos [3], silk worm [4], cricket [5], fruit fly [6,7], zebrafish [8,9], frog [10], mouse [11,12], rat [13], or of human cells [14] this issue presents step-by-step protocols enabling TALEN-assisted mutagenesis for newcomers. For some species we included multiple independent protocols such that interested users have the choice of selecting a procedure following their individual convenience. The TALEN
http://dx.doi.org/10.1016/j.ymeth.2014.08.012 1046-2023/Ó 2014 Elsevier Inc. All rights reserved.
focused horizon of this issue is further widened by the contents of the companion volume edited by C. Giovannangeli and J.-P. Concordet which presents additional applications of TALEN and TAL DNA binding domains and of its successor technology, the CRISP/Cas system [15]. Altogether, TALENs and CRISP/Cas are at the dawn of a new era of reverse genetics now comprising all species and cell lines derived from them. References [1] [2] [3] [4] [5] [6] [7] [8] [9] [10] [11] [12] [13] [14] [15]
K. Chen, Q. Shan, C. Gao, Methods 69 (2014) 2–8. T. Li, B. Liu, C.Y. Chen, B. Yang, Methods 69 (2014) 9–16. A. Aryan, K.M. Myles, Z.N. Adelman, Methods 69 (2014) 38–45. Y. Takasu, T. Tamura, S. Sajwan, I. Kobayashi, M. Zurovec, Methods 69 (2014) 46–57. T. Watanabe, S. Noji, T. Mito, Methods 69 (2014) 17–21. X. Zhang, I.R.S. Ferreira, F. Schnorrer, Methods 69 (2014) 32–37. J. Liu, Y. Chen, R. Jiao, Methods 69 (2014) 22–31. W.Y. Hwang, R.T. Peterson, J.-R.J. Yeh, Methods 69 (2014) 76–84. P. Huang, A. Xiao, X. Tong, Y. Zu, Z. Wang, B. Zhang, Methods 69 (2014) 67–75. Y. Liu, D. Luo, Y. Lei, W. Hu, H. Zhao, C.H.K. Cheng, Methods 69 (2014) 58–66. Y.H. Sung, Y. Jin, S. Kim, H.-W. Lee, Methods 69 (2014) 85–93. B. Wefers, O. Ortiz, W. Wurst, R. Kühn, Methods 69 (2014) 94–101. S. Ménoret, L. Tesson, S. Rémy, C. Usal, V. Thépenier, R. Thinard, et al, Methods 69 (2014) 102–107. Y.-H. Kim, S. Ramakrishna, H. Kim, J.-S. Kim, Methods 69 (2014) 108–117. C. Giovannangeli, Methods (2014), http://dx.doi.org/10.1016/j.ymeth. 2014.08.013.
Ralf Kühn Benedikt Wefers
