Basic Vector Information
pTriEx-mCherry-PA-Rac1 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 ℃.
pTriEx-mCherry-PA-Rac1 vector Sequence
LOCUS 40924_44247 6747 bp DNA circular SYN 13-MAY-2021
DEFINITION synthetic circular DNA.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6747)
AUTHORS Wu YI, Frey D, Lungu OI, Jaehrig A, Schlichting I, Kuhlman B, Hahn
KM
TITLE A genetically encoded photoactivatable Rac controls the motility of
living cells.
JOURNAL Nature. 2009 Sep 3. 461(7260):104-8.
PUBMED 19693014
REFERENCE 2 (bases 1 to 6747)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 6747)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Nature.
2009 Sep 3. 461(7260):104-8."
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..6747
/mol_type="other DNA"
/organism="synthetic DNA construct"
misc_recomb complement(1..706)
/label=baculovirus recombination region (ORF1629)
/note="contains part of ORF1629"
terminator complement(719..766)
/label=T7 terminator
/note="transcription terminator for bacteriophage T7 RNA
polymerase"
primer_bind 836..855
/label=rbglobpA-R
/note="Rabbit beta-globin polyA, reverse primer. Also
called rb-glob-pA-term-R"
polyA_signal complement(854..909)
/label=beta-globin poly(A) signal
/note="rabbit beta-globin polyadenylation signal (Gil and
Proudfoot, 1987)"
primer_bind 955..974
/label=Bglob-pA-R
/note="Rabbit beta-globin polyA region, reverse primer"
CDS complement(1064..1087)
/codon_start=1
/label=8xHis
/note="8xHis affinity tag"
/translation="HHHHHHHH"
CDS complement(1094..1126)
/codon_start=1
/label=HSV tag
/note="HSV (herpes simplex virus) epitope tag"
/translation="QPELAPEDPED"
CDS complement(1166..1738)
/codon_start=1
/label=Rac1 (Q61L)
/note="constitutively active mutant of human Rac family
small GTPase 1"
/translation="ELIKCVVVGDGAVGKTCLLISYTTNAFPGEYIPTVFDNYSANVMV
DGKPVNLGLWDTAGLEDYDRLRPLSYPQTDVFLICFSLVSPASFHHVRAKWYPEVRHHC
PNTPIILVGTKLDLRDDKDTIEKLKEKKLTPITYPQGLAMAKEIGAVKYLECSALTQRG
LKTVFDEAIRAVLCPPPVKKRKRKCLLL"
CDS complement(2168..2872)
/codon_start=1
/label=mCherry
/note="monomeric derivative of DsRed fluorescent protein
(Shaner et al., 2004)"
/translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
GRHSTGGMDELYK"
CDS complement(2879..2896)
/codon_start=1
/label=6xHis
/note="6xHis affinity tag"
/translation="HHHHHH"
promoter complement(2917..3026)
/label=p10 promoter
/note="baculovirus promoter for expression in insect cells"
protein_bind complement(3042..3066)
/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)."
promoter complement(3067..3085)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(3127..3146)
/label=pCAG-F
/note="Rabbit beta-globin intron, for pCAG plasmids,
forward primer"
primer_bind complement(3208..3230)
/label=pCAGGS-5
/note="Chimeric intron in CAG promoter, forward primer"
promoter complement(3436..3635)
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
enhancer complement(3637..3940)
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
misc_recomb complement(4008..4835)
/label=baculovirus recombination region (lef2/ORF603)
/note="contains ORF603 and part of lef2"
CDS 5078..5935
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
rep_origin 5951..6538
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
protein_bind 6706..6739
/label=loxP
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (ATGTATGC) (Shaw et al., 2021)."
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