LOCUS       sequence-227516-        5744 bp    DNA     linear   UNA 14-APR-2025
DEFINITION  CAG-FLEx-iBARK(D110A)-p2A-mCherry: Negative control viral expression
            vector for Cre-dependent Gaq silencing.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 5744)
  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 5744)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 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 D110A p2A mCherry
FEATURES             Location/Qualifiers
     source          1..5744
                     /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    1476..1509
                     /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    1560..1593
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    1618..2206
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1671..1691)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2089..2100)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2249..2370
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2260..2279)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2314..2333
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   2532..2661
                     /label=AAV2 ITR
     rep_origin      complement(2898..3486)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2978..2997)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3657..4448,4449..4517))
                     /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     4280..4299
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4518..4622)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4648..5103)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4785..4806)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4997..5016
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   5175..5304
                     /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 tggcgaagat 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 catggtggtg ctagcataac ttcgtataaa gtatcctata
     1501 cgaagttatt tgccttaacc cagaaattat cactgttatt ctttagaatg gtgcaaagaa
     1561 taacttcgta taatgtatgc tatacgaagt tatgaattcg atatcaagct tatcgataat
     1621 caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct
     1681 tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg
     1741 gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg
     1801 cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt
     1861 tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt
     1921 gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg
     1981 ggcactgaca attccgtggt gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc
     2041 tgtgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat
     2101 ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc
     2161 cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcatcg ataccgtcga
     2221 cccgggcggc cgcttcgagc agacatgata agatacattg atgagtttgg acaaaccaca
     2281 actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat tgctttattt
     2341 gtaaccatta taagctgcaa taaacaagtt aacaacaaca attgcattca ttttatgttt
     2401 caggttcagg gggagatgtg ggaggttttt taaagcaagt aaaacctcta caaatgtggt
     2461 aaaatcgata aggatcttcc tagagcatgg ctacgtagat aagtagcatg gcgggttaat
     2521 cattaactac aaggaacccc tagtgatgga gttggccact ccctctctgc gcgctcgctc
     2581 gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc
     2641 agtgagcgag cgagcgcgca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     2701 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     2761 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     2821 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     2881 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     2941 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     3001 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     3061 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     3121 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     3181 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     3241 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     3301 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct
     3361 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     3421 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     3481 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
     3541 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat
     3601 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac
     3661 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt
     3721 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt
     3781 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag
     3841 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct
     3901 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt
     3961 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc
     4021 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt
     4081 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg
     4141 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg
     4201 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct
     4261 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc
     4321 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt
     4381 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt
     4441 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg
     4501 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat
     4561 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
     4621 cgcacatttc cccgaaaagt gccacctaaa ttgtaagcgt taatattttg ttaaaattcg
     4681 cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc
     4741 cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt tggaacaaga
     4801 gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg
     4861 atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg tgccgtaaag
     4921 cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga aagccggcga
     4981 acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg ctggcaagtg
     5041 tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg ctacagggcg
     5101 cgtcccattc gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt
     5161 cgctattacg ccagctgcgc gctcgctcgc tcactgaggc cgcccgggca aagcccgggc
     5221 gtcgggcgac ctttggtcgc ccggcctcag tgagcgagcg agcgcgcaga gagggagtgg
     5281 ccaactccat cactaggggt tccttgtagt taatgattaa cccgccatgc tacttatcta
     5341 cgtagccatg ctctaggaag agtaccattg acgtcaataa tgacgtatgt tcccatagta
     5401 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
     5461 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
     5521 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
     5581 tacatctacg tattagtcat cgctattacc atggtcgagg tgagccccac gttctgcttc
     5641 actctcccca tctccccccc ctccccaccc ccaattttgt atttatttat tttttaatta
     5701 ttttgtgcag cgatgggggc gggggggggg ggggggcgcg cgcc
//
