Basic Vector Information
- Vector Name:
- pUC57-Simple-gRNA backbone
- Antibiotic Resistance:
- Ampicillin
- Length:
- 2845 bp
- Type:
- CRISPR ; Xenopus expression
- Replication origin:
- ori
- Copy Number:
- High Copy
- Promoter:
- T7
- Cloning Method:
- Restriction Enzyme
- 5' Primer:
- M13R
- 3' Primer:
- M13F
pUC57-Simple-gRNA backbone vector Vector Map
Plasmid Resuspension 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 ℃.
pUC57-Simple-gRNA backbone vector Sequence
LOCUS 40924_45198 2845 bp DNA circular SYN 13-MAY-2021
DEFINITION gRNA expression plasmid.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 2845)
AUTHORS Guo X, Zhang T, Hu Z, Zhang Y, Shi Z, Wang Q, Cui Y, Wang F, Zhao H,
Chen Y
TITLE Efficient RNA/Cas9-mediated genome editing in Xenopus tropicalis.
JOURNAL Development. 2014 Jan 8.
PUBMED 24401372
REFERENCE 2 (bases 1 to 2845)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 2845)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName:
"Development. 2014 Jan 8."
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..2845
/mol_type="other DNA"
/organism="synthetic DNA construct"
protein_bind 55..76
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 91..121
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 129..145
/label=lac operator
/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 153..169
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
promoter 234..252
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
misc_RNA 271..346
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
primer_bind complement(398..414)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(623..642)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 742..764
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind complement(802..820)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 888..992
/label=AmpR promoter
CDS 993..1850
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
rep_origin 2024..2612
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind 2766..2783
/label=L4440
/note="L4440 vector, forward primer"
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