|
Zinc Fingers
|
|
|
|
| 1992–1993 |
Novel DNA triplets |
Structure-based modeling |
(49–51) |
| 1994–1995 |
Novel DNA triplets |
Phage Display |
(52–55) |
| 1999 |
Novel sites with GNN triplets |
|
(56) |
| 2000 |
Novel 9 basepair targets |
Bacterial two-hybrid selections |
(61) |
| 2001 |
Novel triplets with ANN and CNN triplets |
Phage Display |
(57,58) |
| 2001 |
Novel 9 basepair targets |
Phage Display; hybrid 3 finger library panning |
(60) |
| 2001 |
Novel 12 to 18 basepair targets |
Assembly of two-finger ZFP subunits |
(64) |
| 2002 - 2003 |
Drosophia yellow gene target |
Modular assembly |
(31,32) |
| 2005 |
Human IL2Rg gene target |
Zinc finger selections and assembly |
(22) |
| 2008 |
Novel 9 basepair targets |
Bacterial two-hybrid selections |
(62) |
| 2011 |
Novel 9 basepair targets |
Informatics-driven, Context-dependent ZFP assembly |
(63) |
|
Meganucleases
|
|
|
|
| 2002 |
Single basepair target variants (I-CreI) |
Bacterial gene elimination assay/screen |
(97) |
| 2002 |
Activity-based selection (I-SceI) |
Bacterial gene elimination assay/screen |
(98) |
| 2002 |
Hybrid nuclease generation (I-CreI/I-DmoI –> H-DreI |
Structure-based computational redesign |
(110) |
| 2003 |
Single basepair target variants (PI-SceI) |
Bacterial two-hybrid selections |
(96) |
| 2006 |
Single basepair target variant (I-MsoI |
Structure-based computational redesign |
(99) |
| 2006 |
Multiple base pair target variants (I-CreI) |
Bacterial ene elmination assay/screen |
(100) |
| 2006 |
Multiple basepair target variants (I-CreI) |
Eukaryotic gene recombination assay/screen |
(101–103) |
| 2009 |
Individual and multiple base pair target variants (I-AniI) |
Structure-based computational redesign and bacterial selections |
(89) |
| 2009–2010 |
Monomerization of homodimeric meganuclease (I-CreI) |
Structure-based modeling and activity-based selections |
(113,114) |
| 2009 |
Activity-based selections (I-AniI) |
Yeast surface display |
(130) |
| 2010 |
Maize liguless gene target (I-CreI) |
Structure-based modeling and activity-based selections |
(113) |
| 2010 |
Multiple basepair target variants (I-MsoI) |
Structure-based computational redesign |
(105) |
| 2014 |
Human Brutons Tyrosine Kinase (Btk) get target (I-AniI) |
Structure-based computational redesign and Yeast Surface Display |
(107) |
| 2007–2013 |
Various eukaryotic gene targets (I-CreI) |
Structure-based modeling; bacterial selections; eukaryotic selections |
(117–129) |
| 2014–2015 |
Human TCRa and CCR5 gene targets (I-OnuI) |
Yeast surface display meganuclease selections and MegaTAL |
(115,116) |
| 2014–2015 |
Human CFTR gene target (I-OnuI) |
In vitro compartmentalization |
(133,134) |
| 2017 |
Various eukaryotic gene targets (I-OnuI) |
Yeast surface display and bacterial selections |
(23) |
|
TAL effectors
|
|
|
|
| 2009 |
TAL effector code determination and first designer TALs |
Tandem repeat assembly |
(16,136) |
| 2010–2011 |
TAL Nuclease Creation and Initial refinement |
Tandem repeat assembly and FokI fusion |
(171–173) |
| 2012 |
Improved design using additional RVDs; G-specific RVDs |
Tandem repeat assembly using new specificity determinants and data |
(141–143) |
| 2012–2013 |
Increasing mismatch tolerance of C-terminal repeats |
Tand repeat assembly and DNA substrate sequence variation |
(151,152) |
| 2013–2014 |
Altered specificity at base 0 |
Modification of cryptic repeat sequences; RVD at position1, and context |
(132,149,150) |
| 2014 |
Aberrant repeats that allow frameshift binding |
Incorporation of natural repeat variants with small insertions or deletions |
(191) |
| 2014–2015 |
Expanded repertoire of RVDs for fine-tuned targeting |
Characterization of specificities and affinities of 400 RVDs |
(185,186) |
| 2016–2017 |
Modularion of TAL effector binding strength and TALEN efficiency |
Varying the backbone (non-RVD) sequecnes of the repeats |
(188,189) |
| 2017 |
Optimized length for maximum specificity |
Varying the number of repeats |
(153) |
|
Site specific recombinases
|
| 1999 |
Circumvent need for accessory factors (Tn3 resolvase) |
Error prone PCR, galK-based colored colony selection |
(203) |
| 2009 |
Increased efficiency/selectivity (PhiC31 Integrase) |
Error prone PCR, lacZ selection & GFP expression |
(204) |
| 1988 |
Enhanced and altered activity (gin) |
Chemical mutagenesis and bacterial selection |
(205) |
| 2000 |
Circumvent need for accessory factors (lambda integrase) |
GFP based fluorescence |
(206) |
| 2015 |
Targeting CCR5 and AAVS1 safe harbor locus (Bin and Tn21 recombinases) |
Site specific sequence randomization and error prone PCR, antibotic selection |
(207) |
| 2003 |
Altered loxP sequence (Cre) |
site specific sequence randomization, GFP expression and FACS |
(218,219) |
| 2001–2011 |
Altered loxP sequence,HIV LTR sequences (Cre) |
Substrate linked protein evolution |
(207,221,222, 224,226) |
| 2013 |
HIV LTR (improved activity) (Cre) |
Molecular modeling and dynamics |
(227) |
| 2017 |
HIV LTR (improved activity) (Cre) |
Observations based on the crystal structure |
(228) |
| 2008 |
Mutants that promote heterotetramers (Cre) |
Structure-based selection of interfactial residues to be randomized |
(232) |
| 2015 |
Mutants that promote heterotetramers (Cre) |
Protein design via molecular modeling |
(233) |
| 2013 |
Weakened protein–protein interactions to enhance specificity (Cre) |
Random mutagenesis and bacterial selection |
(234) |
| 1988 |
Enhanced activity (Flp) |
Substrate linked protein evolution |
(235) |
| 2004 |
Mutants that promote heterotetramers (Flp) |
Error prone PCR, blue/white selection |
(236) |
| 2003–2006 |
Altered FRT sequence, interleukin 10 target (Flp) |
Error prone PCR and randomization of specific sites, LacZ and RFP reporters |
(237,238) |
| 2016 |
Enhanced activity (R and TD recombinases) |
Sequence truncation, random mutagenesis |
(240) |
| 1995 |
Relaxed specificity (lambda integrase) |
Analysis of chimeric integrases |
(245) |
| 2015 |
Human genome target (lambda integrase) |
Beta-lactamase inhibitor based screen |
(246) |
| 2001 |
Human chromosome 8 target (PhiC31) |
Blue/white selection |
(248) |
| 2003–2011 |
Hybrid reslovase/ZFN targets (serine recombinases) |
Truncated resolvases with zinc finger fusion (varied linkers) |
(249–251) |
| 2011–2014 |
Mutants to promote heterodimers (resolvases) |
Rational design and directed evolution |
(255,256) |
| 2011–2014 |
Altered specificity of catalytic domains (serine recombinases) |
Random mutagenesis of selected residues and directed evolution |
(257,258,261) |
|
Restriction endonucleases
|
| 1987–1999 |
1st Attempts to alter specificity (EcoRI, EcoRV, BamHI) |
Structure-based modeling |
(270–273) |
| 2002–2006 |
Additional attempts to alter specificity (BstYI, NotI) |
Directed evolution and selection |
(274,275) |
| 2003 |
Alteration of specificity of bifunctional RM enzyme (Eco57I) |
Directed evolution and selection for altered methylation specificity |
(278) |
| 2009 |
Alteration of specificity of type IIG enzyme (MmeI) |
Informatics covariation analysis and structure-based modeling |
(21) |
