Basic Vector Information
- Vector Name:
- pMESP23E2
- Antibiotic Resistance:
- Ampicillin
- Length:
- 5601 bp
- Type:
- Cloning vector
- Replication origin:
- ori
- Source/Author:
- Uchanski T, Masiulis S, Fischer B, Kalichuk V
pMESP23E2 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 ℃.
pMESP23E2 vector Sequence
LOCUS 62056_16755 5601 bp DNA circular SYN 07-JAN-2021
DEFINITION Cloning vector pMESP23E2, complete sequence.
ACCESSION MT338521
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5601)
AUTHORS Uchanski T, Masiulis S, Fischer B, Kalichuk V, Lopez-Sanchez U,
Zarkadas E, Weckener M, Sente A, Ward P, Wohlkoenig A, Zoegg T,
Remaut H, Naismith JH, Nury H, Vranken W, Aricescu AR, Pardon E,
Steyaert J.
TITLE Megabodies expand the nanobody toolkit for protein structure
determination by single-particle cryo-EM
JOURNAL Nat Methods 18 (1), 60-68 (2021)
REFERENCE 2 (bases 1 to 5601)
AUTHORS Steyaert J, Uchanski T, Pardon E.
TITLE Direct Submission
JOURNAL Submitted (14-APR-2020) VIB-VUB Center for Structural Biology, VIB,
Pleinlaan 2, Brussels 1050, Belgium
REFERENCE 3 (bases 1 to 5601)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Nat
Methods"; date: "2021"; volume: "18"; issue: "1"; pages: "60-68"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
(14-APR-2020) VIB-VUB Center for Structural Biology, VIB, Pleinlaan
2, Brussels 1050, Belgium"
COMMENT ##Assembly-Data-START##
Sequencing Technology :: Sanger dideoxy sequencing
##Assembly-Data-END##
FEATURES Location/Qualifiers
source 1..5601
/mol_type="other DNA"
/organism="synthetic DNA construct"
protein_bind 107..128
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 143..173
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 181..197
/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)."
sig_peptide 225..290
/label=pelB signal sequence
/note="leader peptide for secretion"
misc_feature 291..326
/label=nanobody beta-strain A (residues 1-13)
/note="nanobody beta-strain A (residues 1-13)"
misc_feature 330..2654
/label=cYgjK scaffold protein
/note="cYgjK scaffold protein"
misc_feature 2658..2690
/label=multi cloning site
/note="multi cloning site"
CDS 2691..2708
/label=6xHis
/note="6xHis affinity tag"
misc_feature 2709..2720
/label=EPEA-tag
/note="EPEA-tag"
primer_bind complement(2730..2746)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
rep_origin 2959..3414
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 3696..3800
/label=AmpR promoter
CDS 3801..4658
/label=AmpR
/note="beta-lactamase"
rep_origin 4832..5420
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
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