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A modular toolbox for gRNA-Cas9 genome engineering in plants based on the GoldenBraid standard.
Vazquez-Vilar M, Bernabe-Orts JM, Fernandez-Del-Carmen A, Ziarsolo P, Blanca J, Granell A, Orzaez D
Plant Methods. 2016 Feb 1;12:10. doi: 10.1186/s13007-016-0101-2. eCollection 2016.
PubMed

Plasmids from Article

ID Plasmid Purpose  
75399pdCas9 (GB1079)Provides the human codon optimized CDS of Cas9 protein with mutated (D10A, H840A) and inactivated catalytic domains as a level 0 GoldenBraid part for C-terminal fusions
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75400pEGB2Ω2_SF-35s:hCas9:tNos (GB1103)Transcriptional unit for (human codon optimized) Cas9 plant expression driven by the 35S promoter. Specially conceived to be linked with the TU of the kanamycin resistance gene GB1181.
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75401pEGB 35s:dCas:BRD:tNos (GB1172)Transcriptional unit of (human codon optimized) inactivated Cas9 fused to the BRD Transcriptional Repressor
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75403pEGB 35s:dCas:VP64:tNos (GB1189)Transcriptional unit of (human codon optimized) inactivated Cas9 fused to the VP64 Transcriptional Activator
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75404pEGB 35s:dCas:EDLL:tNos (GB1190)Transcriptional unit of (human codon optimized) inactivated Cas9 fused to the EDLL Transcriptional Activator
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75405pU6-1 (GB1204)Provides the A. thaliana U6-1 RNA polIII promoter as a level 0 GoldenBraid part
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75406tRNA-gRNA position [D1_2] (GB1205)tRNA and scaffold for the assembly of GBoligomers for the first position (positon [D1_2]) of a polycistronic tRNA-gRNA regulated by the dicot U6-26 or U6-1 promoter (3-part multiplexing)
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75407tRNA-gRNA position [2_n-1] (GB1206)tRNA and scaffold for the assembly of GBoligomers for the intermediate position (positon [2_n-1]) of a polycistronic tRNA-gRNA (3-part multiplexing)
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75408tRNA-gRNA position [n] (GB1207)tRNA and scaffold for the assembly of GBoligomers for the last position (positon [n]) of a polycistronic tRNA-gRNA (2- and 3-part multiplexing)
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75409tRNA-gRNA position [D1_n-1] (GB1208)tRNA and scaffold for the assembly of GBoligomers for the first position (positon [D1_n-1]) of a polycistronic tRNA-gRNA regulated by the U6-26 or U6-1 promoter (2-part multiplexing)
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75410tRNA-gRNA position [M1_2] (GB1209)tRNA and scaffold for the assembly of GBoligomers for the first position (positon [M1_2]) of a polycistronic tRNA-gRNA regulated by the (monocot) U3 promoter (3-part multiplexing)
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75411tRNA-gRNA position [M1_n-1] (GB1210)tRNA and scaffold for the assembly of GBoligomers for the first position (positon [M1_n-1]) of a polycistronic tRNA-gRNA regulated by the (monocot) U3 promoter (2-part multiplexing)
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75412pEGB SF-35S:Renilla:TNOS-35S:P19:TNOS (GB0160)Module for the expression of the Renilla Luciferase with the silencing suppressor P19
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75471pEGB3omega1R Tnos:NptII:Pnos-SF (GB1181)Transcriptional unit for kanamycin resistance gene
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