LOCUS       sequence-339591-        7116 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Cre-dependent GCaMP6s and mBeRFP expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 7116)
  AUTHORS   Melzer S, Newmark ER, Mizuno GO, Hyun M, Philson AC, Quiroli E, 
            Righetti B, Gregory MR, Huang KW, Levasseur J, Tian L, Sabatini BL
  TITLE     Bombesin-like peptide recruits disinhibitory cortical circuits and 
            enhances fear memories.
  JOURNAL   Cell. 2021 Oct 28;184(22):5622-5634.e25. doi: 
            10.1016/j.cell.2021.09.013. Epub 2021 Oct 4.
  PUBMED    34610277
REFERENCE   2  (bases 1 to 7116)
  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.cell.2021.09.013"; journalName: "Cell"; date: 
            "2021-10-28- 28"; volume: "184"; issue: "22"; pages: "5622-5634.e25"
COMMENT     Sequence Label: pAAV-CBA-DIO-GCaMP6s-P2A-mBeRFP-WPRE-pA
FEATURES             Location/Qualifiers
     source          1..7116
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     promoter        251..528
                     /label=chicken beta-actin promoter
     primer_bind     808..830
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     892..911
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    complement(945..978)
                     /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(1029..1062)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(1084..1797)
                     /codon_start=1
                     /product="monomeric derivative of red fluorescent protein 
                     from Entacmaea quadricolor (Merzlyak et al., 2007)"
                     /label=TagRFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTM
                     RIKVVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERSTTYEDGG
                     VLTATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTEMLYPADGGLEGRDY
                     MALKLVGGGHLICNAKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETSVEQHEVAV
                     ARYCDLPSKLGHKL*"
     CDS             complement(1084..1797)
                     /codon_start=1
                     /product="monomeric photoactivatable red fluorescent 
                     protein (Gunewardene et al., 2011)"
                     /label=PAmKate
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTM
                     RIKVVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERSTTYEDGG
                     VLTATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTEMLYPADGGLEGRDY
                     MALKLVGGGHLICNAKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETSVEQHEVAV
                     ARYCDLPSKLGHKL*"
     CDS             complement(1798..1854)
                     /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(join(1864..2304,2305..2310,2311..2745,2746..2763,
                     2764..3033,3034..3039,3040..3096,3097..3123,3124..3147,
                     3148..3150,3151..3183,3184..3186,3187..3204,3205..3213))
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6s
                     /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK
                     TGHAVRAIGRLSSLENVYIKADKQKNGIKANFHIRHNIEDGGVQLAYHYQQNTPIGDGP
                     VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE
                     LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT
                     YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE
                     LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEAFSLFDKDGDGTITTKELGTVMRS
                     LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGN
                     GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     primer_bind     2420..2439
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2679..2700
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2806..2827)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      3108..3117
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      3210..3219
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3238..3271
                     /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    3322..3355
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3368..3956
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3421..3441)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3839..3850)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3999..4120
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4010..4029)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4064..4083
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4282..4411
                     /label=AAV2 ITR
     rep_origin      complement(4648..5236)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4728..4747)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5407..6198,6199..6267))
                     /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     6030..6049
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6268..6372)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6398..6853)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6535..6556)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6747..6766
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   6925..7054
                     /label=AAV2 ITR
ORIGIN
        1 ccattgacgt caataatgac gtatgttccc atagtaacgc caatagggac tttccattga
       61 cgtcaatggg tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat
      121 atgccaagta cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc
      181 cagtacatga ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct
      241 attaccatgg tcgaggtgag ccccacgttc tgcttcactc tccccatctc ccccccctcc
      301 ccacccccaa ttttgtattt atttattttt taattatttt gtgcagcgat gggggcgggg
      361 gggggggggg ggcgcgcgcc aggcggggcg gggcggggcg aggggcgggg cggggcgagg
      421 cggagaggtg cggcggcagc caatcagagc ggcgcgctcc gaaagtttcc ttttatggcg
      481 aggcggcggc ggcggcggcc ctataaaaag cgaagcgcgc ggcgggcggg agtcgctgcg
      541 acgctgcctt cgccccgtgc cccgctccgc cgccgcctcg cgccgcccgc cccggctctg
      601 actgaccgcg ttactcccac aggtgagcgg gcgggacggc ccttctcctc cgggctgtaa
      661 ttagcgcttg gtttaatgac ggcttgtttc ttttctgtgg ctgcgtgaaa gccttgaggg
      721 gctccgggag ggccctttgt gcggggggag cggctcgggg ctgtccgcgg ggggacggct
      781 gccttcgggg gggacggggc agggcggggt tcggcttctg gcgtgtgacc ggcggctcta
      841 gagcctctgc taaccatgtt catgccttct tctttttcct acagctcctg ggcaacgtgc
      901 tggttattgt gctgtctcat cattttggca aagaattgga tcttataact tcgtatagga
      961 tactttatac gaagttatgc agaatggtag ctggattgta gctgctatta gcaatatgaa
     1021 acctcttaat aacttcgtat agcatacatt atacgaagtt atactagtgc ggccgcggaa
     1081 ttcttattaa agtttgtgcc ccagtttgct agggaggtcg cagtatctgg ccacagccac
     1141 ctcgtgctgc tcgacggagg tctctttgtc ggcctccttg attctttcca gtcttctgtc
     1201 cacatagtag acgccgggca tcttgaggtt cttagcgggt ttcttggatc tgtatgtggt
     1261 cttggcgttg cagatcaggt ggcccccgcc cacgagcttc agggccatgt agtctctgcc
     1321 ttccaggccg ccgtcagcgg ggtacagcat ctcggtgctg gcctcccagc cgagtgtttt
     1381 cttctgcatc acagggccgt tggatgggaa gttcacccct ctgatcttga cgttgtagat
     1441 gaggcagccg tcctggaggc tggtgtcctg ggtagcggtc agcacgcccc cgtcttcgta
     1501 tgtggtggat ctctcccatg tgaagccctc agggaaggac tgcttaaaga agtcggggat
     1561 gccctgggtg tggttgatga aggtcttgct gccgtacatg aagctggtag ccaggatgtc
     1621 gaaggcgaag gggagagggc cgccctcgac caccttgatt ctcatggtct gggtgccctc
     1681 gtagggcttg ccttcgccct cggatgtgca cttgaagtgg tggttgttca cggtgccctc
     1741 catgtacagc ttcatgtgca tgttctcctt aatcagctct tcgcccttag acaccatagg
     1801 accggggttt tcttccacgt ctcctgcttg ctttaacaga gagaagttcg tggctccgga
     1861 tcccttcgct gtcatcattt gtacaaactc ttcgtagttt acctgaccat ccccatcgat
     1921 gtctgcttcc ctgatcattt catcaacctc ttcatctgtt aacttctctc caaggtttgt
     1981 catcacgtgg cgaagctctg ctgcactgat gtagccattg ccatccttat caaacacacc
     2041 gaacgcttct ctaatttctt cttccgtgtc cctgtatttc atttttcttg ccatcattgt
     2101 caggaactca gggaagtcga ttgtgccgtc accgtcggca tctacttcat tgatcatgtc
     2161 ctgcagctct gcttctgtgg ggttctgccc cagagaccgc atcaccgtcc ccagctcctt
     2221 ggttgttatt gtcccatccc cgtccttgtc aaatagggag aaagcctctt taaattctgc
     2281 gatctgctct tcagtcagtt ggtccggcag gttgtactcc agcttgtgcc ccaggatgtt
     2341 gccgtcctcc ttgaagtcga tgcccttcag ctcgatgcgg ttcaccaggg tgtcgccctc
     2401 gaacttcacc tcggcgcggg tcttgtagtt gccgtcgtcc ttgaagaaga tggtgcgctc
     2461 ctggatgtag ccttcgggca tggcggactt gaagaagtcg tgctgcttca tgtggtcggg
     2521 gtagcggctg aagcactgca cgccgtaggt cagggtggtc acgagggtgg gccagggcac
     2581 gggcagcttg ccggtggtgc agatgaactt cagggtcagc ttgccgtagg tggcatcgcc
     2641 ctcaccctcg ccggacacgc tgaacttgtg gccgtttacg tcgccgtcca gctcgaccag
     2701 gatgggcacc accccggtga acagctcctc gcccttgctc accatgctcc ctccggtacc
     2761 gcccttgtac agctcgtcca tgccgagagt gatcccggcg gcggtcacga actccagcag
     2821 gaccatgtga tcgcgcttct cgttggggtc tttcgaaagt ttggactgca cgctcaggta
     2881 gtggttgtcg ggcagcagca cggggccgtc gccgatgggg gtgttctgct ggtagtggta
     2941 ggcgagctgc acgccgccgt cctcgatgtt gtggcggatg tggaagttcg ccttgatgcc
     3001 gttcttctgc ttgtcggcct tgatatagac gttctcgagt gagctcagcc gacctatagc
     3061 tctgactgcg tgacctgtct tattccactt acgacgtgat gagtcgacca tggtggcgag
     3121 atccttatcg tcatcgtcgt acagatcccg acccatttgc tgtccaccag tcatgctagc
     3181 cataccatga tgatgatgat gatgagaacc catggtggca ccggtactag tctcgagata
     3241 acttcgtata aagtatccta tacgaagtta tttgccttaa cccagaaatt atcactgtta
     3301 ttctttagaa tggtgcaaag aataacttcg tataatgtat gctatacgaa gttataagct
     3361 tatcgataat caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta
     3421 tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc
     3481 ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga
     3541 ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac
     3601 ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc
     3661 cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc
     3721 tcggctgttg ggcactgaca attccgtggt gttgtcgggg aaatcatcgt cctttccttg
     3781 gctgctcgcc tgtgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc
     3841 ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc
     3901 gcgtcttcgc cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcatcg
     3961 ataccgtcga cccgggcggc cgcttcgagc agacatgata agatacattg atgagtttgg
     4021 acaaaccaca actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat
     4081 tgctttattt gtaaccatta taagctgcaa taaacaagtt aacaacaaca attgcattca
     4141 ttttatgttt caggttcagg gggagatgtg ggaggttttt taaagcaagt aaaacctcta
     4201 caaatgtggt aaaatcgata aggatcttcc tagagcatgg ctacgtagat aagtagcatg
     4261 gcgggttaat cattaactac aaggaacccc tagtgatgga gttggccact ccctctctgc
     4321 gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc
     4381 gggcggcctc agtgagcgag cgagcgcgca gctgcattaa tgaatcggcc aacgcgcggg
     4441 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     4501 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     4561 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     4621 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     4681 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     4741 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     4801 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     4861 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     4921 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     4981 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     5041 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     5101 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     5161 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     5221 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     5281 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     5341 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     5401 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     5461 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     5521 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     5581 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     5641 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     5701 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     5761 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     5821 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     5881 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     5941 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     6001 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     6061 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     6121 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     6181 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     6241 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     6301 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     6361 aggggttccg cgcacatttc cccgaaaagt gccacctaaa ttgtaagcgt taatattttg
     6421 ttaaaattcg cgttaaattt ttgttaaatc agctcatttt ttaaccaata ggccgaaatc
     6481 ggcaaaatcc cttataaatc aaaagaatag accgagatag ggttgagtgt tgttccagtt
     6541 tggaacaaga gtccactatt aaagaacgtg gactccaacg tcaaagggcg aaaaaccgtc
     6601 tatcagggcg atggcccact acgtgaacca tcaccctaat caagtttttt ggggtcgagg
     6661 tgccgtaaag cactaaatcg gaaccctaaa gggagccccc gatttagagc ttgacgggga
     6721 aagccggcga acgtggcgag aaaggaaggg aagaaagcga aaggagcggg cgctagggcg
     6781 ctggcaagtg tagcggtcac gctgcgcgta accaccacac ccgccgcgct taatgcgccg
     6841 ctacagggcg cgtcccattc gccattcagg ctgcgcaact gttgggaagg gcgatcggtg
     6901 cgggcctctt cgctattacg ccagctgcgc gctcgctcgc tcactgaggc cgcccgggca
     6961 aagcccgggc gtcgggcgac ctttggtcgc ccggcctcag tgagcgagcg agcgcgcaga
     7021 gagggagtgg ccaactccat cactaggggt tccttgtagt taatgattaa cccgccatgc
     7081 tacttatcta cgtagccatg ctctaggaag atcgta
//
