Basic Vector Information
- Vector Name:
- AAV-P(Per2)-DIO-intron2-dLUC
- Antibiotic Resistance:
- Ampicillin
- Length:
- 7917 bp
- Type:
- Mammalian Expression, AAV, Cre/Lox
- Replication origin:
- ori
- Copy Number:
- High Copy
- Promoter:
- Per2
- Cloning Method:
- Gibson Cloning
AAV-P(Per2)-DIO-intron2-dLUC 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 ℃.
AAV-P(Per2)-DIO-intron2-dLUC vector Sequence
LOCUS 40924_155 7917 bp DNA circular SYN 13-MAY-2021
DEFINITION Per2 transcription luciferase reporter.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7917)
AUTHORS Mei L, Fan Y, Lv X, Welsh DK, Zhan C, Zhang EE
TITLE Long-term in vivo recording of circadian rhythms in brains of freely
moving mice.
JOURNAL Proc Natl Acad Sci U S A. 2018 Apr 2. pii: 1717735115. doi:
10.1073/pnas.1717735115.
PUBMED 29610316
REFERENCE 2 (bases 1 to 7917)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 7917)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Proc Natl
Acad Sci U S A. 2018 Apr 2. pii: 1717735115. doi:
10.1073/pnas.1717735115."
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..7917
/mol_type="other DNA"
/organism="synthetic DNA construct"
repeat_region 1..130
/label=AAV2 ITR (alternate)
/note="Functional equivalent of wild-type AAV2 ITR"
protein_bind complement(689..722)
/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)."
protein_bind complement(773..806)
/label=loxP
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (ATGTATGC) (Shaw et al., 2021)."
CDS complement(812..931)
/codon_start=1
/label=hPEST
/note="PEST degradation sequence from mouse ornithine
decarboxylase"
/translation="SHGFPPEVEEQAAGTLPMSCAQESGMDRHPAACASARINV"
CDS complement(935..982)
/codon_start=1
/label=hCL1
/note="non-ORF yeast peptide conferring ubiquitin-dependent
degradation"
/translation="ACKNWFSSLSHFVIHL"
CDS complement(989..2635)
/codon_start=1
/label=luciferase
/note="firefly luciferase"
/translation="EDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDAH
IEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAPA
NDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQSM
YTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHAR
DPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYKI
QSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGYG
LTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMSG
YVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESILL
QHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVVF
VDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
CDS complement(2645..2665)
/codon_start=1
/label=TEV site
/note="tobacco etch virus (TEV) protease recognition and
cleavage site"
/translation="ENLYFQG"
CDS complement(2690..2734)
/codon_start=1
/label=AviTag(TM)
/note="peptide tag that allows for enzymatic biotinylation"
/translation="GLNDIFEAQKIEWHE"
CDS complement(2756..2821)
/codon_start=1
/label=3xFLAG
/note="three tandem FLAG(R) epitope tags, followed by an
enterokinase cleavage site"
/translation="DYKDHDGDYKDHDIDYKDDDDK"
CDS complement(3548..3571)
/codon_start=1
/label=FLAG
/note="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
/translation="DYKDDDDK"
regulatory 3571..3580
/label=Kozak sequence
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
/regulatory_class="other"
promoter complement(3618..3636)
/label=SP6 promoter
/note="promoter for bacteriophage SP6 RNA polymerase"
promoter complement(3642..3660)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
intron complement(3705..3837)
/label=chimeric intron
/note="chimera between introns from human beta-globin and
immunoglobulin heavy chain genes"
protein_bind 3902..3935
/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)."
protein_bind 3986..4019
/label=loxP
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (ATGTATGC) (Shaw et al., 2021)."
misc_feature 4044..4632
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
polyA_signal 4664..5140
/label=hGH poly(A) signal
/note="human growth hormone polyadenylation signal"
repeat_region 5180..5320
/label=AAV2 ITR
/note="inverted terminal repeat of adeno-associated virus
serotype 2"
rep_origin 5395..5850
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
primer_bind complement(5867..5886)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
primer_bind 5986..6008
/label=pGEX 3'
/note="pGEX vectors, reverse primer"
primer_bind complement(6046..6064)
/label=pBRforEco
/note="pBR322 vectors, upsteam of EcoRI site, forward
primer"
promoter 6132..6236
/label=AmpR promoter
CDS 6237..7094
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
rep_origin 7268..7856
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
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