Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V010644 | pRN227 | In stock, 1 week for quality controls |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
- Vector Name:
- pRN227
- Antibiotic Resistance:
- Kanamycin
- Length:
- 20260 bp
- Type:
- Plant Expression
- Replication origin:
- ori
- Copy Number:
- High Copy
- Promoter:
- CaMV35S(enhanced)
- Cloning Method:
- Restriction Enzyme
pRN227 vector Map
Plasmid Protocol
1. Centrifuge at 5,000×g for 5 min.
2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.
3. Close the tube and incubate for 10 minutes at room temperature.
4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.
5. Store the plasmid at -20 ℃.
6. The concentration of plasmid re-measurement sometimes differs from the nominal value, which may be due to the position of the lyophilized plasmid in the tube, the efficiency of the re-dissolution, the measurement bias, and adsorption on the wall of the tube, therefore, it is recommended to transform and extract the plasmid before using it
General Plasmid Transform Protocol
1. Take one 100μl of the competent cells and thaw it on ice for 10min, add 2μl of plasmid, then ice bath for 30min, then heat-shock it at 42℃ for 60s, do not stir, and then ice bath for 2min.
2. Add 900μl of LB liquid medium without antibiotics, and incubate at 37℃ for 45min (30℃ for 1-1.5 hours) with 180rpm shaking.
3. Centrifuge at 6000rpm for 5min, leave only 100μl of supernatant to resuspend the bacterial precipitate and spread it onto the target plasmid-resistant LB plate.
4. Invert the plate and incubate at 37℃ for 14h, or at 30℃ for 20h.
5. Pick a single colony into LB liquid medium, add the corresponding antibiotics, incubate at 220rpm for 14h, and extract the plasmid according to the experimental needs and the instructions of the plasmid extraction kit.
pRN227 vector Sequence
LOCUS V010644 20260 bp DNA circular SYN 13-MAY-2021
DEFINITION Exported.
ACCESSION V010644
VERSION V010644
KEYWORDS pRN227
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
.
REFERENCE 1 (bases 1 to 20260)
AUTHORS Maher MF, Nasti RA, Vollbrecht M, Starker CG, Clark MD, Voytas DF
TITLE Plant gene editing through de novo induction of meristems.
JOURNAL Nat Biotechnol. 2019 Dec 16. pii: 10.1038/s41587-019-0337-2. doi:
10.1038/s41587-019-0337-2.
PUBMED 31844292
REFERENCE 2 (bases 1 to 20260)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 20260)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Nat
Biotechnol. 2019 Dec 16. pii: 10.1038/s41587-019-0337-2. doi:
10.1038/s41587-019-0337-2."
SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..20260
/mol_type="other DNA"
/organism="synthetic DNA construct"
misc_feature 1..25
/label="LB T-DNA repeat"
/note="left border repeat from nopaline C58 T-DNA"
polyA_signal 103..277
/label="CaMV poly(A) signal"
/note="cauliflower mosaic virus polyadenylation signal"
CDS complement(337..1125)
/label="NeoR/KanR"
/note="aminoglycoside phosphotransferase"
promoter complement(1194..1870)
/label="CaMV 35S promoter (enhanced)"
/note="cauliflower mosaic virus 35S promoter with a
duplicated enhancer region"
protein_bind 2061..2082
/label="CAP binding site"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 2097..2127
/label="lac promoter"
/note="promoter for the E. coli lac operon"
protein_bind 2135..2151
/label="lac operator"
/bound_moiety="lac repressor encoded by lacI"
/note="The lac repressor binds to the lac operator to
inhibit transcription in E. coli. This inhibition can be
relieved by adding lactose or
isopropyl-beta-D-thiogalactopyranoside (IPTG)."
primer_bind 2140..2162
/label="M13/pUC Reverse"
/note="In lacZ gene"
primer_bind 2159..2175
/label="M13 rev"
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 2159..2175
/label="M13 Reverse"
/note="In lacZ gene. Also called M13-rev"
promoter 3080..3425
/label="CaMV 35S promoter"
/note="strong constitutive promoter from cauliflower mosaic
virus"
CDS 3459..7562
/label="Cas9"
/note="Cas9 (Csn1) endonuclease from the Streptococcus
pyogenes Type II CRISPR/Cas system"
CDS 7575..7595
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label="SV40 NLS"
/translation="PKKKRKV"
terminator 7608..7856
/label="HSP terminator"
/note="efficient transcription terminator from the
Arabidopsis thaliana heat shock protein 18.2 gene (Nagaya
et al., 2010)"
promoter 7904..8327
/label="AtU6-26"
/note="Arabidopsis U6-26 gene promoter"
misc_RNA 8349..8424
/label="gRNA scaffold"
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
promoter 8559..8742
/label="NOS promoter"
/note="nopaline synthase promoter"
CDS 10970..12115
/gene="STM"
/label="Homeobox protein SHOOT MERISTEMLESS"
/note="Homeobox protein SHOOT MERISTEMLESS from Arabidopsis
thaliana. Accession#: Q38874"
polyA_signal 12125..12299
/label="CaMV poly(A) signal"
/note="cauliflower mosaic virus polyadenylation signal"
CDS complement(12732..13607)
/note="Replication-associated protein A from Bean yellow
dwarf virus. Accession#: O39521"
/label="Replication-associated protein A"
misc_feature 13981..14005
/label="RB T-DNA repeat"
/note="right border repeat from nopaline C58 T-DNA"
CDS 15306..15932
/label="pVS1 StaA"
/note="stability protein from the Pseudomonas plasmid pVS1
(Heeb et al., 2000)"
CDS 16364..17434
/label="pVS1 RepA"
/note="replication protein from the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
rep_origin 17503..17697
/label="pVS1 oriV"
/note="origin of replication for the Pseudomonas plasmid
pVS1 (Heeb et al., 2000)"
primer_bind complement(17933..17955)
/label="pGEX 3'"
/note="pGEX vectors, reverse primer"
misc_feature 18041..18181
/label="bom"
/note="basis of mobility region from pBR322"
primer_bind complement(18196..18213)
/label="L4440"
/note="L4440 vector, forward primer"
rep_origin complement(18367..18955)
/direction=LEFT
/label="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(19045..19836)
/label="KanR"
/note="aminoglycoside phosphotransferase"