Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V000033 | pDEST22 | 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:
- pDEST22
- Antibiotic Resistance:
- Ampicillin
- Length:
- 8930 bp
- Type:
- Yeast Two-Hybrid System Vectors
- Replication origin:
- ori
- Host:
- Yeast
- Promoter:
- ADH1(long)
pDEST22 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.
pDEST22 vector Sequence
LOCUS 40924_14440 8930 bp DNA circular SYN 13-JAN-2022
DEFINITION synthetic circular DNA.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 8930)
TITLE Direct Submission
REFERENCE 2 (bases 1 to 8930)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..8930
/mol_type="other DNA"
/organism="synthetic DNA construct"
protein_bind 81..102
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 117..147
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 155..171
/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 179..195
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
promoter 216..234
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
promoter 1018..1722
/label=ADH1 promoter
/note="promoter for alcohol dehydrogenase 1"
CDS 1734..1754
/codon_start=1
/label=SV40 NLS
/note="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/translation="PKKKRKV"
CDS 1764..2105
/codon_start=1
/label=GAL4 activation domain
/note="activation domain of the GAL4 transcriptional
activator"
/translation="ANFNQSGNIADSSLSFTFTNSSNGPNLITTQTNSQALSQPIASSN
VHDNFMNNEITASKIDDGNNSKPLSPGWTDQTAYNAFGITTGMFNTTTMDDVYNYLFDD
EDTPPNPKKE"
protein_bind 2121..2245
/label=attR1
/note="recombination site for the Gateway(R) LR reaction"
promoter 2392..2494
/label=cat promoter
/note="promoter of the E. coli cat gene encoding
chloramphenicol acetyltransferase"
CDS 2495..3172
/codon_start=1
/label=CmR
/note="chloramphenicol acetyltransferase"
/translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQAGRNLE
DPAY"
CDS 3495..3797
/codon_start=1
/label=ccdB
/note="CcdB, a bacterial toxin that poisons DNA gyrase"
/translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
VPRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
protein_bind complement(3841..3965)
/label=attR2
/note="recombination site for the Gateway(R) LR reaction"
terminator 4165..4352
/label=ADH1 terminator
/note="transcription terminator for the S. cerevisiae
alcohol dehydrogenase 1 (ADH1) gene"
promoter complement(4505..4523)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(4530..4546)
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
rep_origin 4687..5142
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
CDS complement(5248..5919)
/codon_start=1
/label=TRP1
/note="phosphoribosylanthranilate isomerase, required for
tryptophan biosynthesis"
/translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
KK"
promoter complement(5920..6201)
/label=TRP1 promoter
misc_feature complement(6459..6962)
/label=CEN/ARS
/note="S. cerevisiae CEN6 centromere fused to an
autonomously replicating sequence"
promoter 6999..7103
/label=AmpR promoter
CDS 7104..7961
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
rep_origin 8135..8723
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"