pet28-MBP-CBE-nC9-His F576
Ligation of Cas9-His* into F350pet28-MBP-CBE-nC9*
- Gene 2
- CBE
- Other markers
- kan\sequence
- Size
- 11,980 bp, circular
- Record date
- 01-JUN-2023
Construct
- T7 promoter–MBP–ORF-1–GamApo_String1–GAM–APOBEC1–GamApo_String2–nCas9 · payload cassette, 20–6,832
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 |
|---|---|---|---|
| XbaI | T^CTAGA | 66 | between lac\operator and MBP |
| NcoI | C^CATGG | 105 | between lac\operator and MBP |
| BsaI (Eco31I) | GGTCTC(1/5) | 233 | in MBP |
| BsiWI (Pfl23II) | C^GTACG | 460 | in MBP |
| SalI | G^TCGAC | 1,353 | in ORF-1 |
| SacII (Cfr42I) | CCGC^GG | 1,717 | in GAM |
| AatII (ZraI) | GACGT^C | 2,684 | in SGGS Linker' |
| BamHI | G^GATCC | 2,726 | in ORF-1 |
| SwaI (SmiI) | ATTT^AAAT | 3,249 | in nCas9' |
| NheI (BmtI) | G^CTAGC | 3,827 | between ORF-1 and 'nCas9 |
| XhoI (PaeR7I) | C^TCGAG | 6,854 | between ORF-1 and ORF1 |
| SmaI (XmaI, TspMI) | CCC^GGG | 8,085 | in kan\sequence |
| ClaI (BspDI) | AT^CGAT | 8,266 | in kan\sequence |
| SapI (BspQI, LguI) | GCTCTTC(1/4) | 9,274 | between kan\sequence and lac\I |
| ApaI (PspOMI) | GGGCC^C | 11,055 | in lac\I |
Cut twice: HindIII (3,735, 6,080), SphI (3,989, 11,791), KpnI (1,926, 4,506), BglII (529, 11,980), MluI (5,300, 11,258), BstBI (1,884, 4,066), NsiI (7,847, 8,113), PvuI (495, 7,961).
No site: NotI, SpeI, AscI, PacI, SfiI, SbfI, FseI, PmeI, AvrII, AgeI, StuI, AflII, BsrGI, EagI, 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 |
|---|---|---|---|---|---|
| T7 promoter | Site CDS | 20 | 37 | 18 | + |
| lac\operator | Site CDS | 36 | 66 | 31 | + |
| MBP | Coding sequence CDS | 107 | 1,353 | 1,247 | + |
| ORF-1 | Coding sequence CDS | 107 | 3,827 | 3,721 | + |
| 6xHis affinity tag | Site CDS | 119 | 136 | 18 | + |
| thrombin recognition and cleavage site | Site CDS | 146 | 163 | 18 | + |
| 'GamApo_String1 | Coding sequence CDS | 1,354 | 2,054 | 701 | + |
| TEV | Site misc_feature | 1,361 | 1,381 | 21 | + |
| GAM | Coding sequence CDS | 1,382 | 1,952 | 571 | + |
| APOBEC1 | Coding sequence CDS | 1,953 | 2,636 | 684 | + |
| GamApo_String2 | Coding sequence CDS | 2,025 | 2,726 | 702 | + |
| SGGS Linker' | Coding sequence CDS | 2,636 | 2,726 | 91 | + |
| nCas9' | Coding sequence CDS | 2,733 | 3,827 | 1,095 | + |
| D10A | Site misc_feature | 2,756 | 2,758 | 3 | + |
| ''nCas9 | Coding sequence CDS | 3,828 | 6,831 | 3,004 | + |
| ORF-1 | Coding sequence CDS | 3,828 | 6,832 | 3,005 | + |
| 'ORF1 | Site CDS | 6,855 | 6,880 | 26 | + |
| His-Tag | Site CDS | 6,860 | 6,877 | 18 | + |
| Stop | Site CDS | 6,878 | 6,880 | 3 | + |
| kan\sequence | Selection marker CDS | 7,579 | 8,391 | 813 | − |
| lac\I | Coding sequence CDS | 10,534 | 11,613 | 1,080 | − |
PromoterCoding sequenceTerminatorSelection markerOrigin / backboneT-DNA borderSite
Feature types are inferred from the labels in the GenBank file; the file itself is the reference.