Basic Vector Information
- Vector Name:
- pUDE215
- Antibiotic Resistance:
- Ampicillin
- Length:
- 11766 bp
- Type:
- Expression vector
- Replication origin:
- ori
- Source/Author:
- Kozak BU, van Rossum HM, Benjamin KR, Wu L, Daran JM, Pronk JT, van Maris AJ.
- Promoter:
- TPI1
pUDE215 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 ℃.
pUDE215 vector Sequence
LOCUS 40924_45463 11766 bp DNA circular SYN 18-DEC-2018
DEFINITION Expression vector pUDE215, complete sequence.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 11766)
AUTHORS Kozak BU, van Rossum HM, Benjamin KR, Wu L, Daran JM, Pronk JT, van
Maris AJ.
TITLE Replacement of the Saccharomyces cerevisiae acetyl-CoA synthetases
by alternative pathways for cytosolic acetyl-CoA synthesis
JOURNAL Metab. Eng. (2013) In press
PUBMED 24269999
REFERENCE 2 (bases 1 to 11766)
AUTHORS Kozak BU, van Rossum HM, Benjamin KR, Wu L, Daran J-M.G., Pronk JT,
van Maris AJA.
TITLE Direct Submission
JOURNAL Submitted (31-MAY-2013) Biotechnology, Delft University of
Technology, Julianalaan 67, Delft 2628BC, The Netherlands
REFERENCE 3 (bases 1 to 11766)
TITLE Direct Submission
REFERENCE 4 (bases 1 to 11766)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Metab. Eng.
(2013) In press"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
(31-MAY-2013) Biotechnology, Delft University of Technology,
Julianalaan 67, Delft 2628BC, The Netherlands"
COMMENT SGRef: number: 3; type: "Journal Article"
COMMENT ##Assembly-Data-START##
Assembly Method :: IDBA v. 0.20
Sequencing Technology :: Illumina
##Assembly-Data-END##
FEATURES Location/Qualifiers
source 1..11766
/mol_type="other DNA"
/organism="synthetic DNA construct"
rep_origin complement(183..771)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(945..1802)
/label=AmpR
/note="beta-lactamase"
promoter complement(1803..1907)
/label=AmpR promoter
rep_origin complement(1934..3276)
/direction=LEFT
/label=2u ori
/note="yeast 2u plasmid origin of replication"
promoter 3538..3753
/label=URA3 promoter
CDS 3754..4554
/label=URA3
/note="orotidine-5'-phosphate decarboxylase, required for
uracil biosynthesis"
rep_origin complement(4688..5143)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind 5284..5300
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
promoter 5310..5328
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
promoter 5838..6357
/label=TPI1 promoter
/note="strong constitutive promoter for yeast triose
phosphate isomerase"
CDS 6358..8646
/codon_start=1
/gene="PFL"
/product="pyruvate formate lyase"
/label=PFL
/protein_id="AHB33291.1"
/translation="MTLKASKKAAVPVMQAKTYATAPVITNDAAAKSEIDVEGWIKKHY
TPYEGDGSFLAGPTEKTKKLFAKAEEYLAKERANGGLYDVDPHTPSTITSHKPGYLDKE
NEVIYGYQTDVPLKRAIKPFGGVNMVKNALKAVNVPMDKEVEHIFSDYRKTHNTAVFDI
YSKEMRAGRSNAIMTGLPDGYGRGRIIGDYRRVALYGTDRLIAQKEKDKAELQKKQMDE
PTMKLIGEVADQVKALKQLTQMAKSYGIDITKPAKNAREATQFVYFGYLGSIKEQDGAA
MSLGRVDAFLDCFFENDLKNGVITESEAQEIIDNLILKLRFARHLRTPEYNDLFAGDPT
WVTMSLGGMGSDGRTLVTKTSFRVLNTLYNLGPAPEPNITVLWNKNLPKNFKDFATKVS
IDTSSIQYESDALMSARFGDDYGIACCVSAMRIGKDMQFFGARCNLAKLMLYVLNHGKD
ERTGKQVGPDFGPVPEGPIPFDWMWETYDKAMDWIAKLYVNTMNVIHFCHDQYCYESLQ
MALHDTDVRRLMAFGVAGLSVVADSFSAIKYAKVTPIRDPKTGLTVDFKVEGEFPKFGN
DDDRVDFFARTVTDKLINKLRKTPTYRGATHTLSILTITSNVVYGKKTGSTPDGRKAGQ
PFAPGCNPMHGREFSGAVASLSSVAKVNYDSCMDGISNTFSIVPNTIGKTLQERQGNLS
GLLDGYFSKGAHHLNVNVLKRETLEDAMAHPENYPNLTIRVSGYAVNFVKLTPAQQKEV
IARTFHEKM"
gene 6358..8646
/gene="PFL"
/label=PFL
regulatory 8647..9147
/label=GND2 terminator
/note="GND2 terminator"
/regulatory_class="terminator"
regulatory 9152..10152
/label=FBA1 promoter
/note="FBA1 promoter"
/regulatory_class="promoter"
CDS 10153..10953
/codon_start=1
/gene="PFL-AE"
/product="pyruvate formate lyase activating enzyme"
/label=PFL-AE
/protein_id="AHB33292.1"
/translation="MPAIVDPTTMDYMEVKGNVHSTESLACLEGPGNRFLLFLNGCAAR
CLYCSNPDTWDETVGTPMTVGQLIKKIGNLKNYYINSVGGGGVTVSGGEPLTQFGFLSC
FLYAVKKHLNLHTCVETTGQGCTKAWNSVLPHTDLCLVCIKHAIPEKYEQITRTKKLDR
CLKFLKELEKRNIPWWCRYVVLPGYTDSKEDIEALIELVKNSPTCERIEFLPYPELGKN
KWEELGIEYPLKNIKQLKKSEIKWICDMVREAFKDRNIPVTGDT"
gene 10153..10953
/gene="PFL-AE"
/label=PFL-AE
regulatory 10954..11454
/label=PMA1 terminator
/note="PMA1 terminator"
/regulatory_class="terminator"
primer_bind complement(11455..11471)
/label=SK primer
/note="common sequencing primer, one of multiple similar
variants"
promoter complement(11508..11526)
/label=T3 promoter
/note="promoter for bacteriophage T3 RNA polymerase"
primer_bind complement(11547..11563)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind 11571..11587
/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)."
promoter complement(11595..11625)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind complement(11640..11661)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
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