Basic Vector Information
| Basic Vector Information | |||
|---|---|---|---|
| Vector Name | pTpPuc3 | Antibiotic Resistance | Kanamycin |
| Length | 7796 bp | Type | Cloning vector |
| Source | Karas BJ, Diner RE, Lefebvre SC, Weyman PD. | ||
pTpPuc3 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 ℃.
pTpPuc3 vector Sequence
LOCUS Exported 7796 bp ds-DNA circular SYN 18-DEC-2018
DEFINITION Cloning vector pTpPuc3, complete sequence.
ACCESSION KP745603
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7796)
AUTHORS Karas BJ, Diner RE, Lefebvre SC, Weyman PD.
TITLE Designer Diatom episomes Delivered by Bacterial Conjugation
JOURNAL Unpublished
REFERENCE 2 (bases 1 to 7796)
AUTHORS Karas BJ, Diner RE, Lefebvre SC, Weyman PD.
TITLE Direct Submission
JOURNAL Submitted (03-FEB-2015) Synthetic Biology and Bioenergy Group, J.
Craig Venter Institute, 4120 Capricorn Lane, La Jolla, CA 92037, USA
REFERENCE 3 (bases 1 to 7796)
AUTHORS .
TITLE Direct Submission
COMMENT ##Assembly-Data-START##
Sequencing Technology :: Sanger dideoxy sequencing
##Assembly-Data-END##
FEATURES Location/Qualifiers
source 1..7796
/organism="Cloning vector pTpPuc3"
/mol_type="other DNA"
/label=episome vector
/note="episome vector"
/db_xref="taxon:1655162"
misc_feature 185..956
/label=OriT from RP4
/note="OriT from RP4"
CDS complement(307..678)
/codon_start=1
/gene="traJ"
/product="oriT-recognizing protein"
/label=traJ
/translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
KQDELGKVMMGVVRPRAEP"
oriT 711..820
/note="incP origin of transfer"
primer_bind 1098..1114
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
misc_feature 1116..3178
/note="Nourseothricin resistance (Nat) cassette for T.
pseudonana"
CDS 2096..2665
/codon_start=1
/gene="Streptomyces noursei nat1"
/product="nourseothricin acetyltransferase"
/label=NrsR
/note="confers resistance to nourseothricin"
/translation="MTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGFT
LREVPVDPPLTKVFPDDESDDESDAGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNRR
LTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFTL
CGLDTALYDGTASDGEQALYMSMPCP"
misc_feature 3179..4561
/label=yeast Cen6-ArsH4-His3
/note="yeast Cen6-ArsH4-His3"
misc_feature 3185..3688
/label=CEN/ARS
/note="S. cerevisiae CEN6 centromere fused to an
autonomously replicating sequence"
promoter 3689..3876
/gene="S. cerevisiae HIS3"
/label=HIS3 promoter
CDS 3877..4536
/codon_start=1
/gene="S. cerevisiae HIS3"
/product="imidazoleglycerol-phosphate dehydratase, required
for histidine biosynthesis"
/label=HIS3
/note="yeast auxotrophic marker"
/translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
KEALLARGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHFL
ESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
primer_bind complement(4605..4621)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind 4629..4645
/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(4653..4683)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 4698..4719
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
misc_feature complement(5007..5595)
/label=MB1 origin of replication
/note="MB1 origin of replication"
rep_origin complement(5007..5595)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
misc_feature 6320..7289
/label=KanR gene from ACYC177
/note="KanR gene from ACYC177"
CDS 6443..7258
/codon_start=1
/gene="aph(3')-Ia"
/product="aminoglycoside phosphotransferase"
/label=KanR
/note="confers resistance to kanamycin in bacteria or G418
(Geneticin(R)) in eukaryotes"
/translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
promoter complement(7597..7701)
/gene="bla"
/label=AmpR promoter
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