title: Optimized tuning of TALEN specificity using non-conventional RVDs.
abstract: A key feature when designing DNA targeting tools and especially nucleases is specificity. The ability to control and tune this important parameter represents an invaluable advance to the development of such molecular scissors. Here, we identified and characterized new non-conventional RVDs (ncRVDs) that possess novel intrinsic targeting specificity features. We further report a strategy to control TALEN targeting based on the exclusion capacities of ncRVDs (discrimination between different nucleotides). By implementing such ncRVDs, we demonstrated in living cells the possibility to efficiently promote TALEN-mediated processing of a target in the HBB locus and alleviate undesired off-site cleavage. We anticipate that this method can greatly benefit to designer nucleases, especially for therapeutic applications and synthetic biology.
authors: CELLECTIS S.A., 8 Rue de la Croix Jarry, 75013 Paris, France.
authors: Alexandre
authors: A
authors: Juillerat
authors: CELLECTIS S.A., 8 Rue de la Croix Jarry, 75013 Paris, France.
authors: Coline
authors: C
authors: Pessereau
authors: CELLECTIS S.A., 8 Rue de la Croix Jarry, 75013 Paris, France.
authors: Gwendoline
authors: G
authors: Dubois
authors: CELLECTIS S.A., 8 Rue de la Croix Jarry, 75013 Paris, France.
authors: Valérie
authors: V
authors: Guyot
authors: CELLECTIS S.A., 8 Rue de la Croix Jarry, 75013 Paris, France.
authors: Alan
authors: A
authors: Maréchal
authors: CELLECTIS S.A., 8 Rue de la Croix Jarry, 75013 Paris, France.
authors: Julien
authors: J
authors: Valton
authors: CELLECTIS S.A., 8 Rue de la Croix Jarry, 75013 Paris, France.
authors: Fayza
authors: F
authors: Daboussi
authors: CELLECTIS S.A., 8 Rue de la Croix Jarry, 75013 Paris, France.
authors: Laurent
authors: L
authors: Poirot
authors: CELLECTIS S.A., 8 Rue de la Croix Jarry, 75013 Paris, France.
authors: Aymeric
authors: A
authors: Duclert
authors: CELLECTIS S.A., 8 Rue de la Croix Jarry, 75013 Paris, France.
authors: Philippe
authors: P
authors: Duchateau
mesh_terms: D001483:Base Sequence; D002460:Cell Line; D003851:Deoxyribonucleases / Q000378:metabolism*; D018900:Dinucleotide Repeats; D018390:Gene Targeting; D006454:Hemoglobins / Q000737:chemistry / Q000235:genetics; D006801:Humans; D006868:Hydrolysis; D008969:Molecular Sequence Data; D013379:Substrate Specificity; D015003:Yeasts / Q000235:genetics / Q000378:metabolism
chemical_list: D006454:Hemoglobins; D003851:Deoxyribonucleases
references: Nat Biotechnol. 2011 Aug;29(8):699-700
references: 21822242
references: Science. 2012 Feb 10;335(6069):716-9
references: 22223736
references: Science. 2012 Feb 10;335(6069):720-3
references: 22223738
references: Nat Biotechnol. 2012 May;30(5):460-5
references: 22484455
references: Nat Biotechnol. 2012 Jul;30(7):593-5
references: 22781676
references: Nucleic Acids Res. 2012 Jul;40(13):6367-79
references: 22467209
references: Nat Commun. 2012;3:968
references: 22828628
references: Nucleic Acids Res. 2012 Aug;40(15):7584-95
references: 22581776
references: PLoS One. 2012;7(9):e45383
references: 23028976
references: J Biol Chem. 2012 Nov 9;287(46):38427-32
references: 23019344
references: Nucleic Acids Res. 2012 Nov;40(21):11163-72
references: 23019222
references: Nat Biotechnol. 2013 Jan;31(1):76-81
references: 23242165
references: Nucleic Acids Res. 2013 Apr;41(7):4118-28
references: 23408851
references: Nat Commun. 2013;4:1762
references: 23612303
references: Biotechnol Bioeng. 2013 Jul;110(7):1811-21
references: 23508559
references: Nature. 2013 Aug 22;500(7463):472-6
references: 23877069
references: Acta Crystallogr D Biol Crystallogr. 2013 Sep;69(Pt 9):1707-16
references: 23999294
references: Nat Biotechnol. 2013 Sep;31(9):833-8
references: 23907171
references: Curr Gene Ther. 2013 Aug;13(4):291-303
references: 23888878
references: Nucleic Acids Res. 2013 Oct;41(19):9197-207
references: 23921635
references: Nat Struct Mol Biol. 2013 Nov;20(11):1321-4
references: 24096363
references: Nat Biotechnol. 2013 Dec;31(12):1133-6
references: 24013198
references: Nat Biotechnol. 2013 Dec;31(12):1137-42
references: 24108092
references: PLoS One. 2013;8(12):e82539
references: 24349308
references: Nucleic Acids Res. 2014 Feb;42(4):2591-601
references: 24285304
references: Nat Methods. 2014 Apr;11(4):429-35
references: 24531420
references: Nucleic Acids Res. 2014 Apr;42(8):5390-402
references: 24569350
references: Cell Res. 2014 May;24(5):628-31
references: 24513857
references: Nat Commun. 2014;5:3831
references: 24871200
references: Expert Opin Ther Targets. 2014 Aug;18(8):835-9
references: 24917359
references: BMC Mol Biol. 2014;15:13
references: 24997498
references: Mol Biotechnol. 2014 Aug;56(8):681-8
references: 24870618
references: ACS Synth Biol. 2014 Oct 17;3(10):723-30
references: 24251925
references: J Mol Biol. 2006 Jan 20;355(3):443-58
references: 16310802
references: Science. 2009 Dec 11;326(5959):1501
references: 19933106
references: Science. 2009 Dec 11;326(5959):1509-12
references: 19933107
references: Nat Biotechnol. 2011 Feb;29(2):149-53
references: 21248753
references: Nucleic Acids Res. 2011 Jul;39(12):e82
references: 21493687