Table 1. MS2-CRISPR/dCas9 expression constructs used in this study.
| Plasmid | Description | Effector domain | Effector function |
|---|---|---|---|
| pJL005 |
35S:MS2-GFP:tRBCS 35S:dCas9:tRBCS sgRNA-FT-A sgRNA-FT-B 35S:BastaR:t35S |
GFP | Fluorescent marker (negative control) |
| pMH211 |
35S:MS2-VP64:tRBCS 35S:dCas9:tRBCS sgRNA-FT-A sgRNA-FT-B pMAS:BastaR:tMAS |
VP64 | Transcriptional activator |
| pMH212 |
35S:MS2-VP64:tRBCS pSUC2:dCas9:tRBCS sgRNA-FT-A sgRNA-FT-B pMAS:BastaR:tMAS |
||
| pMH213 |
pSUC2:MS2-VP64:tRBCS 35S:dCas9:tRBCS sgRNA-FT-A sgRNA-FT-B pMAS:BastaR:tMAS |
||
| pMH214 |
pSUC2:MS2-VP64:tRBCS pSUC2:dCas9:tRBCS sgRNA-FT-A sgRNA-FT-B pMAS:BastaR:tMAS |
||
| pJL011 |
35S:MS2-SRDX:tRBCS 35S:dCas9:tRBCS sgRNA-FT-A sgRNA-FT-B pMAS:BastaR:tMAS |
SRDX | Transcriptional repressor |
| pJL044 |
35S:MS2-p300:tRBCS 35S:dCas9:tRBCS sgRNA-FT-A sgRNA-FT-B pMAS:BastaR:tMAS |
Homo sapiens p300 core domain | H3K27 acetyltransferase |
| pJL049 |
35S:MS2-G9a:tRBCS 35S:dCas9:tRBCS sgRNA-FT-A sgRNA-FT-B pMAS:BastaR:tMAS |
Homo sapiens G9a SET domain | H3K9 methyltransferase |
| pJL050 |
35S:MS2-KYP:tRBCS 35S:dCas9:tRBCS sgRNA-FT-A sgRNA-FT-B pMAS:BastaR:tMAS |
Arabidopsis KYP SET domain | H3K9 methyltransferase |
All coding sequences originating from non-plant organisms were codon-optimised for Arabidopsis. tRBCS = terminator of Rubisco gene (from pea); BastaR = Basta resistance gene; t35S = terminator of CaMV 35S; pMAS = promoter of mannopine synthase; tMAS = terminator of mannopine synthase.