Basic Vector Information
- Vector Name:
- DBH-p2a-FLPo
- Antibiotic Resistance:
- Ampicillin
- Length:
- 7156 bp
- Type:
- Cloning vector
- Replication origin:
- p15A ori
- Source/Author:
- Sun JJ, Ray R.
- Promoter:
- mPGK
DBH-p2a-FLPo 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 ℃.
DBH-p2a-FLPo vector Sequence
LOCUS 40924_575 7156 bp DNA circular SYN 17-DEC-2018
DEFINITION Cloning vector DBH-p2a-FLPo, complete sequence.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7156)
AUTHORS Sun JJ, Ray R.
TITLE Generation of Two Noradrenergic-Specific
Dopamine-Beta-Hydroxylase-FLPo Knock-In Mice Using
CRISPR/Cas9-Mediated Targeting in Embryonic Stem Cells
JOURNAL PLoS ONE 11 (7), E0159474 (2016)
PUBMED 27441631
REFERENCE 2 (bases 1 to 7156)
AUTHORS Sun JJ, Ray R.
TITLE Direct Submission
JOURNAL Submitted (27-APR-2016) Neuroscience, Baylor College of Medicine,
One Baylor Plaza Room T707, Houston, TX 77030, USA
REFERENCE 3 (bases 1 to 7156)
TITLE Direct Submission
REFERENCE 4 (bases 1 to 7156)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "PLoS ONE";
date: "2016"; volume: "11"; issue: "7"; pages: "E0159474"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
(27-APR-2016) Neuroscience, Baylor College of Medicine, One Baylor
Plaza Room T707, Houston, TX 77030, USA"
COMMENT SGRef: number: 3; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..7156
/mol_type="other DNA"
/organism="synthetic DNA construct"
rep_origin 18..562
/label=p15A ori
/note="Plasmids containing the medium-copy-number p15A
origin of replication can be propagated in E. coli cells
that contain a second plasmid with the ColE1 origin."
misc_feature 836..1835
/label=5' homology arm
/note="5' homology arm"
CDS 1845..1901
/codon_start=1
/label=P2A
/note="2A peptide from porcine teschovirus-1 polyprotein"
/translation="ATNFSLLKQAGDVEENPGP"
CDS 1902..3197
/codon_start=1
/label=FLPo
/note="nuclear-targeted site-specific recombinase"
/translation="MAPKKKRKVMSQFDILCKTPPKVLVRQFVERFERPSGEKIASCAA
ELTYLCWMITHNGTAIKRATFMSYNTIISNSLSFDIVNKSLQFKYKTQKATILEASLKK
LIPAWEFTIIPYNGQKHQSDITDIVSSLQLQFESSEEADKGNSHSKKMLKALLSEGESI
WEITEKILNSFEYTSRFTKTKTLYQFLFLATFINCGRFSDIKNVDPKSFKLVQNKYLGV
IIQCLVTETKTSVSRHIYFFSARGRIDPLVYLDEFLRNSEPVLKRVNRTGNSSSNKQEY
QLLKDNLVRSYNKALKKNAPYPIFAIKNGPKSHIGRHLMTSFLSMKGLTELTNVVGNWS
DKRASAVARTTYTHQITAIPDHYFALVSRYYAYDPISKEMIALKDETNPIEEWQHIEQL
KGSAEGSIRYPAWNGIISQEVLDYLSSYINRRI"
protein_bind 3201..3234
/label=lox2272
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are
compatible with each other, but incompatible with loxP or
loxN sites (Lee and Saito, 1988)."
promoter 3241..3740
/label=PGK promoter
/note="mouse phosphoglycerate kinase 1 promoter"
promoter 3752..3799
/label=EM7 promoter
/note="synthetic bacterial promoter"
CDS 3818..4618
/codon_start=1
/label=NeoR/KanR
/note="aminoglycoside phosphotransferase from Tn5"
/translation="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
EHQGLAPAELFARLKARMPDGDDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
polyA_signal 4659..4883
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
misc_feature 4938..4971
/label=Lox2722
/note="Lox2722"
protein_bind complement(4938..4971)
/label=lox2272
/bound_moiety="Cre recombinase"
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (GGATACTT). lox2272 sites are compatible with each
other, but incompatible with loxP or loxN sites."
misc_feature 4972..5968
/label=3' homology arm with PAM motif deleted
/note="3' homology arm with PAM motif deleted"
promoter 6044..6148
/label=AmpR promoter
CDS 6149..7006
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
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