LOCUS       sequence-227801-        5759 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  CAG-FLEx-iBARK-p2A-mCherry: Viral expression vector for 
            Cre-dependent Gaq silencing.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 5759)
  AUTHORS   Nagai J, Bellafard A, Qu Z, Yu X, Ollivier M, Gangwani MR, 
            Diaz-Castro B, Coppola G, Schumacher SM, Golshani P, Gradinaru V, 
            Khakh BS
  TITLE     Specific and behaviorally consequential astrocyte Gq GPCR signaling 
            attenuation in vivo with ibetaARK.
  JOURNAL   Neuron. 2021 Jul 21;109(14):2256-2274.e9. doi: 
            10.1016/j.neuron.2021.05.023. Epub 2021 Jun 16.
  PUBMED    34139149
REFERENCE   2  (bases 1 to 5759)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1016/j.neuron.2021.05.023"; journalName: "Neuron"; date: 
            "2021-07-21- 21"; volume: "109"; issue: "14"; pages: "2256-2274.e9"
COMMENT     Sequence Label: CAG FLEX BARK p2A mCherry
FEATURES             Location/Qualifiers
     source          1..5759
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     8..27
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    complement(84..117)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /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(168..201)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(210..920)
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(488..507)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     757..775
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             complement(921..977)
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             complement(1410..1433)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             complement(1434..1460)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     protein_bind    1491..1524
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /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    1575..1608
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    1633..2221
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1686..1706)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2104..2115)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2264..2385
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2275..2294)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2329..2348
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   2547..2676
                     /label=AAV2 ITR
     rep_origin      complement(2913..3501)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2993..3012)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3672..4463,4464..4532))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4295..4314
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4533..4637)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4663..5118)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4800..4821)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5012..5031
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   5190..5319
                     /label=AAV2 ITR
ORIGIN
        1 ctcctgggca acgtgctggt tattgtgctg tctcatcatt ttggcaaaga attggatcgg
       61 gatcctctag agtcgactcc ggaataactt cgtataggat actttatacg aagttatgca
      121 gaatggtagc tggattgtag ctgctattag caatatgaaa cctcttaata acttcgtata
      181 gcatacatta tacgaagtta tggcgcgccc tacttgtaca gctcgtccat gccgccggtg
      241 gagtggcggc cctcggcgcg ttcgtactgt tccacgatgg tgtagtcctc gttgtgggag
      301 gtgatgtcca acttgatgtt gacgttgtag gcgccgggca gctgcacggg cttcttggcc
      361 ttgtaggtgg tcttgacctc agcgtcgtag tggccgccgt ccttcagctt cagcctctgc
      421 ttgatctcgc ccttcagggc gccgtcctcg gggtacatcc gctcggagga ggcctcccag
      481 cccatggtct tcttctgcat tacggggccg tcggagggga agttggtgcc gcgcagcttc
      541 accttgtaga tgaactcgcc gtcctgcagg gaggagtcct gggtcacggt caccacgccg
      601 ccgtcctcga agttcatcac gcgctcccac ttgaagccct cggggaagga cagcttcaag
      661 tagtcgggga tgtcggcggg gtgcttcacg taggccttgg agccgtacat gaactgaggg
      721 gacaggatgt cccaggcgaa gggcaggggg ccacccttgg tcaccttcag cttggcggtc
      781 tgggtgccct cgtaggggcg gccctcgccc tcgccctcga tctcgaactc gtggccgttc
      841 acggagccct ccatgtgcac cttgaagcgc atgaactcct tgatgatggc catgttatcc
      901 tcctcgccct tgctcaccat cggtccagga ttctcttcga catctccggc ttgtttcagc
      961 agagagaagt ttgttgcgcc actaccgtgg atgttgagct ctacattctt ccactggcaa
     1021 aatcgtgtga atttatcgct ctcgatgaat ttctggaaca cgtctcctcg gaggttctgg
     1081 caaatttctt caatatatgg ctggaagaga tccggaggca cctgcttctt caccagatgg
     1141 ccctggacgt gctcaatggc gctcttcgag aaaggatgtg agcaggccag cagctccttc
     1201 atgatgtacg tgtcgaagat ctctcggctg cagaccaggc gctcctcctc tgtctccagc
     1261 ttctcgtatt tcttgatctc ctcgtagaac tctaccaagg gcttggcctc ctccaggtgc
     1321 ttcaggcaga agtctcggaa aagcaggtac cccagcttct gggagaagat cttctcaaaa
     1381 gtcacctcgc cccggtcctc caggtacttc ttgtcgtcat cgtctttgta gtccgcatag
     1441 tcaggaacat cgtatgggta catggtggta agcttgggtc tcccgctagc ataacttcgt
     1501 ataaagtatc ctatacgaag ttatttgcct taacccagaa attatcactg ttattcttta
     1561 gaatggtgca aagaataact tcgtataatg tatgctatac gaagttatga attcgatatc
     1621 aagcttatcg ataatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt
     1681 aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct
     1741 attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt
     1801 tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac
     1861 gcaaccccca ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct
     1921 ttccccctcc ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca
     1981 ggggctcggc tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt
     2041 ccttggctgc tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc
     2101 ccttcggccc tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct
     2161 cttccgcgtc ttcgccttcg ccctcagacg agtcggatct ccctttgggc cgcctccccg
     2221 catcgatacc gtcgacccgg gcggccgctt cgagcagaca tgataagata cattgatgag
     2281 tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat
     2341 gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc
     2401 attcatttta tgtttcaggt tcagggggag atgtgggagg ttttttaaag caagtaaaac
     2461 ctctacaaat gtggtaaaat cgataaggat cttcctagag catggctacg tagataagta
     2521 gcatggcggg ttaatcatta actacaagga acccctagtg atggagttgg ccactccctc
     2581 tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt
     2641 tgcccgggcg gcctcagtga gcgagcgagc gcgcagctgc attaatgaat cggccaacgc
     2701 gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg
     2761 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     2821 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     2881 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     2941 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     3001 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     3061 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     3121 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     3181 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     3241 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     3301 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta
     3361 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     3421 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     3481 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     3541 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     3601 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     3661 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     3721 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     3781 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta
     3841 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     3901 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     3961 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     4021 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     4081 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     4141 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     4201 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     4261 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     4321 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     4381 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     4441 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     4501 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc
     4561 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa
     4621 caaatagggg ttccgcgcac atttccccga aaagtgccac ctaaattgta agcgttaata
     4681 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     4741 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc
     4801 cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa
     4861 ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt
     4921 cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac
     4981 ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta
     5041 gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg
     5101 cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg caactgttgg gaagggcgat
     5161 cggtgcgggc ctcttcgcta ttacgccagc tgcgcgctcg ctcgctcact gaggccgccc
     5221 gggcaaagcc cgggcgtcgg gcgacctttg gtcgcccggc ctcagtgagc gagcgagcgc
     5281 gcagagaggg agtggccaac tccatcacta ggggttcctt gtagttaatg attaacccgc
     5341 catgctactt atctacgtag ccatgctcta ggaagagtac cattgacgtc aataatgacg
     5401 tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt ggagtattta
     5461 cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac gccccctatt
     5521 gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac cttatgggac
     5581 tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt cgaggtgagc
     5641 cccacgttct gcttcactct ccccatctcc cccccctccc cacccccaat tttgtattta
     5701 tttatttttt aattattttg tgcagcgatg ggggcggggg gggggggggg gcgcgcgcc
//
