p3oR-U10Cwt F587
Ligation of gent-Xho-Mlu* into C035p7ioR-Uq10CasWToi*
- Promoter 1
- Ubq10
- Gene 1
- Spec
- Other markers
- Sm/Sp
- Size
- 14,274 bp, circular
- Record date
- 04-JUL-2023
Construct
- 35S–ORF-1 · E9 · payload cassette, 309–1,579
- Pro-Ubq10At–Cas9WT · NOS-T · payload cassette, 1,628–7,580
- SfiI ×2 · flanks the Pro-Ubq10At–Cas9WT cassette · at 1,620, 8,159
- BsmBI Golden Gate · overhangs TGAT, GTTT · at 8,029, 8,061
- T-DNA borders · LB, RB
Read out of the GenBank file; compare every vector in the multiple expression cassettes and cloning sites table.
Files
Order
Vectors from the list cost 250 €, Cas9 vectors 400 €, net. Send the vector name to info@dna-cloning.com and we ship on invoice.
Restriction sites
| Unique cutter | Site | Cuts at | Location |
|---|---|---|---|
| MfeI (MunI) | C^AATTG | 14 | in LB |
| XhoI (PaeR7I) | C^TCGAG | 727 | between 35S and ORF-1 |
| MluI | A^CGCGT | 1,279 | between ORF-1 and E9 |
| PstI | CTGCA^G | 1,589 | between E9 and Pro-Ubq10At |
| SbfI (SdaI) | CCTGCA^GG | 1,589 | between E9 and Pro-Ubq10At |
| SpeI | A^CTAGT | 3,104 | between Pro-Ubq10At and Cas9WT |
| BamHI | G^GATCC | 3,110 | between Pro-Ubq10At and Cas9WT |
| SmaI (XmaI, TspMI) | CCC^GGG | 7,324 | between Cas9WT and NOS-T |
| NsiI (Mph1103I) | ATGCA^T | 7,447 | in NOS-T |
| BstBI (Bsp119I) | TT^CGAA | 8,042 | in targetRNA |
| HindIII | A^AGCTT | 8,045 | in targetRNA |
| ApaI (PspOMI) | GGGCC^C | 8,176 | between RNA' and RB |
| KpnI (Acc65I) | GGTAC^C | 8,182 | between RNA' and RB |
| PmeI (MssI) | GTTT^AAAC | 8,214 | between RNA' and RB |
| FseI | GGCCGG^CC | 11,412 | in pVS1 |
| BsiWI (Pfl23II) | C^GTACG | 11,640 | in pVS1 |
| NheI (BmtI) | G^CTAGC | 11,756 | in pVS1 |
| ClaI (BspDI) | AT^CGAT | 11,836 | in pVS1 |
| BstEII (Eco91I) | G^GTNACC | 13,121 | in Sm/Sp |
Cut twice: EcoRI (2,288, 7,584), SalI (2,466, 8,145), SacI (2,721, 3,166), NcoI (1,615, 4,589), NdeI (9,720, 10,300), SfiI (1,620, 8,159), AgeI (12,250, 12,558), StuI (5,886, 8,153), AatII (3,708, 8,605), BsaI (12,159, 12,579), BsmBI (8,029, 8,061).
No site: AscI, PacI, SwaI, AvrII, HpaI, BsrGI, AarI.
Computed from the GenBank sequence for 50 common enzymes; positions are the cut in the top strand, without Dam/Dcm methylation. Compare vectors in the restriction site finder.
Annotated features
| Feature | Type | Start | End | Length | Strand |
|---|---|---|---|---|---|
| LB | T-DNA border misc_feature | 1 | 24 | 24 | + |
| 35S | Promoter misc_feature | 309 | 724 | 416 | + |
| ORF-1 | Coding sequence CDS | 745 | 1,278 | 534 | + |
| sp | Site misc_feature | 958 | 961 | 4 | + |
| 'E9 | Terminator misc_feature | 1,289 | 1,579 | 291 | + |
| Pro-Ubq10At | Promoter misc_feature | 1,628 | 3,103 | 1,476 | + |
| Ubi cds 5` | Coding sequence CDS | 2,732 | 2,795 | 64 | + |
| íntron | Other misc_feature | 2,798 | 3,101 | 304 | + |
| 'Cas9WT | Coding sequence CDS | 3,125 | 7,321 | 4,197 | + |
| NLS | Site misc_feature | 3,131 | 3,151 | 21 | + |
| NLS | Site misc_feature | 7,271 | 7,318 | 48 | + |
| NOS-T | Terminator misc_feature | 7,327 | 7,580 | 254 | + |
| 'RNA' | Other misc_feature | 7,585 | 8,145 | 561 | + |
| U6At | Coding sequence CDS | 7,590 | 8,034 | 445 | + |
| targetRNA | Site CDS | 8,035 | 8,061 | 27 | + |
| hairpin | Site CDS | 8,062 | 8,091 | 30 | + |
| gRNA | Site misc_feature | 8,062 | 8,144 | 83 | + |
| RB | T-DNA border misc_feature | 8,230 | 8,253 | 24 | + |
| ColE1 | Origin / backbone misc_feature | 8,788 | 9,485 | 698 | + |
| 'pVS1 | Origin / backbone misc_feature | 9,773 | 12,560 | 2,788 | + |
| Sm/Sp | Selection marker CDS | 12,744 | 13,754 | 1,011 | + |
PromoterCoding sequenceTerminatorSelection markerOrigin / backboneT-DNA borderSite
Feature types are inferred from the labels in the GenBank file; the file itself is the reference.