LOCUS       sequence-151076-        7459 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescent reporter for calcium signaling.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7459)
  AUTHORS   Liberti WA 3rd, Markowitz JE, Perkins LN, Liberti DC, Leman DP, 
            Guitchounts G, Velho T, Kotton DN, Lois C, Gardner TJ
  TITLE     Unstable neurons underlie a stable learned behavior.
  JOURNAL   Nat Neurosci. 2016 Oct 10. doi: 10.1038/nn.4405.
  PUBMED    27723744
REFERENCE   2  (bases 1 to 7459)
  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"; journalName: "Nat 
            Neurosci. 2016 Oct 10. doi: 10.1038/nn.4405."
COMMENT     Sequence Label: pHAGE-RSV-GCAMP6f
FEATURES             Location/Qualifiers
     source          1..7459
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             1..634
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    681..806
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1303..1536
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1721..1765
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2058..2175
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2218..2479
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     regulatory      2483..2492
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2489..2497,2498..2515,2516..2518,2519..2551,2552..2554,
                     2555..2578,2579..2605,2606..2662,2663..2668,2669..2938,
                     2939..2956,2957..3391,3392..3397,3398..3841)
                     /codon_start=1
                     /product="improved fluorescent protein-based calcium sensor
                     (Chen et al., 2013)"
                     /label=GCaMP6f
                     /translation="MGSHHHHHHGMASMTGGQQMGRDLYDDDDKDLATMVDSSRRKWNK
                     TGHAVRAIGRLSSLENVYIKADKQKNGIKANFKIRHNIEDGGVQLAYHYQQNTPIGDGP
                     VLLPDNHYLSVQSKLSKDPNEKRDHMVLLEFVTAAGITLGMDELYKGGTGGSMVSKGEE
                     LFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKFICTTGKLPVPWPTLVTTLT
                     YGVQCFSRYPDHMKQHDFFKSAMPEGYIQERTIFFKDDGNYKTRAEVKFEGDTLVNRIE
                     LKGIDFKEDGNILGHKLEYNLPDQLTEEQIAEFKEEFSLFDKDGDGTITTKELGTVMRS
                     LGQNPTEAELQDMINEVDADGDGTIDFPEFLTMMARKMKYRDTEEEIREAFGVFDKDGN
                     GYISAAELRHVMTNLGEKLTDEEVDEMIREADIDGDGQVNYEEFVQMMTAK"
     regulatory      2585..2594
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     2875..2896
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(3002..3023)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3263..3282)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     misc_feature    3849..4437
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3902..3922)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4320..4331)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             4512..4745
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(4864..4882)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4950..5054
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5055..5123,5124..5915)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(5273..5292)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6086..6674
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6575..6594
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6828..6845
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7085..7106
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7121..7126,7127..7144,7145..7151)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7159..7175
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     7164..7186
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7183..7199
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7183..7199
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     7249..7268
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 tggaagggct aattcactcc caaagaagac aagatatcct tgatctgtgg atctaccaca
       61 cacaaggcta cttccctgat tagcagaact acacaccagg gccaggggtc agatatccac
      121 tgacctttgg atggtgctac aagctagtac cagttgagcc agataaggta gaagaggcca
      181 ataaaggaga gaacaccagc ttgttacacc ctgtgagcct gcatgggatg gatgacccgg
      241 agagagaagt gttagagtgg aggtttgaca gccgcctagc atttcatcac gtggcccgag
      301 agctgcatcc ggagtacttc aagaactgct gatatcgagc ttgctacaag ggactttccg
      361 ctggggactt tccagggagg cgtggcctgg gcgggactgg ggagtggcga gccctcagat
      421 cctgcatata agcagctgct ttttgcctgt actgggtctc tctggttaga ccagatctga
      481 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct
      541 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc
      601 agaccctttt agtcagtgtg gaaaatctct agcagtggcg cccgaacagg gacttgaaag
      661 cgaaagggaa accagaggag ctctctcgac gcaggactcg gcttgctgaa gcgcgcacgg
      721 caagaggcga ggggcggcga ctggtgagta cgccaaaaat tttgactagc ggaggctaga
      781 aggagagaga tgggtgcgag agcgtcagta ttaagcgggg gagaattaga tcgcgatggg
      841 aaaaaattcg gttaaggcca gggggaaaga aaaaatataa attaaaacat atagtatggg
      901 caagcaggga gctagaacga ttcgcagtta atcctggcct gttagaaaca tcagaaggct
      961 gtagacaaat actgggacag ctacaaccat cccttcagac aggatcagaa gaacttagat
     1021 cattatataa tacagtagca accctctatt gtgtgcatca aaggatagag ataaaagaca
     1081 ccaaggaagc tttagacaag atagaggaag agcaaaacaa aagtaagacc accgcacagc
     1141 aagcggccgg ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag
     1201 tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc
     1261 aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg
     1321 gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc
     1381 cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc
     1441 gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct
     1501 ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa
     1561 actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca
     1621 gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt
     1681 aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt
     1741 ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta
     1801 tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt
     1861 actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct
     1921 cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga
     1981 cagagacaga tccattcgat tagtgaacgg atctcgacgg tatcgccgaa ttcacaaatg
     2041 gcagtattca tccacaattt taaaagaaaa ggggggattg gggggtacag tgcaggggaa
     2101 agaatagtag acataatagc aacagacata caaactaaag aattacaaaa acaaattaca
     2161 aaaattcaaa attttcgggt ttattacagg gacagcagag atccagtttg gactagtaat
     2221 gtagtcttat gcaatactct tgtagtcttg caacatggta acgatgagtt agcaacatgc
     2281 cttacaagga gagaaaaagc accgtgcatg ccgattggtg gaagtaaggt ggtacgatcg
     2341 tgccttatta ggaaggcaac agacgggtct gacatggatt ggacgaacca ctgaattccg
     2401 cattgcagag atattgtatt taagtgccta gctcgataca ataaacgcca tttgaccatt
     2461 caccacattg gtgtgcaccg cggccgccat gggttctcat catcatcatc atcatggtat
     2521 ggctagcatg actggtggac agcaaatggg tcgggatctg tacgacgatg acgataagga
     2581 tctcgccacc atggtcgact catcacgtcg taagtggaat aagacaggtc acgcagtcag
     2641 agctataggt cggctgagct cactcgagaa cgtctatatc aaggccgaca agcagaagaa
     2701 cggcatcaag gcgaacttca agatccgcca caacatcgag gacggcggcg tgcagctcgc
     2761 ctaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca
     2821 ctacctgagc gtgcagtcca aactttcgaa agaccccaac gagaagcgcg atcacatggt
     2881 cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaaggg
     2941 cggtaccgga gggagcatgg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat
     3001 cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggtga
     3061 gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccaccg gcaagctgcc
     3121 cgtgccctgg cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta
     3181 ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacatcca
     3241 ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt
     3301 cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg
     3361 caacatcctg gggcacaagc tggagtacaa cctgccggac caactgactg aagagcagat
     3421 cgcagaattt aaagaggaat tctccctatt tgacaaggac ggggatggga caataacaac
     3481 caaggagctg gggacggtga tgcggtctct ggggcagaac cccacagaag cagagctgca
     3541 ggacatgatc aatgaagtag atgccgacgg tgacggcaca atcgacttcc ctgagttcct
     3601 gacaatgatg gcaagaaaaa tgaaatacag ggacacggaa gaagaaatta gagaagcgtt
     3661 cggtgtgttt gataaggatg gcaatggcta catcagtgca gcagagcttc gccacgtgat
     3721 gacaaacctt ggagagaagt taacagatga agaggttgat gaaatgatca gggaagcaga
     3781 catcgatggg gatggtcagg taaactacga agagtttgta caaatgatga cagcgaagtg
     3841 aggatcctaa tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact
     3901 atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg
     3961 cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg
     4021 aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa
     4081 cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc
     4141 ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg
     4201 ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt
     4261 ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt
     4321 cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc
     4381 cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcctg
     4441 agatccttta agaccaatga cttacaaggc agctgtagat cttagccact ttttaaaaga
     4501 aaagggggga ctggaagggc taattcactc ccaacgaaga caagatctgc tttttgcttg
     4561 tactgggtct ctctggttag accagatctg agcctgggag ctctctggct aactagggaa
     4621 cccactgctt aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct
     4681 gttgtgtgac tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc
     4741 tagcagtagt agttcatgtc atcttattat tcagtattta taacttgcaa agaaatgaat
     4801 atcagagagt gagaggcccg ggttaattaa ggaaagggct agatcattct tgaagacgaa
     4861 agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga
     4921 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     4981 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     5041 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     5101 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     5161 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     5221 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     5281 gcgcggtatt atcccgtgtt gacgccgggc aagagcaact cggtcgccgc atacactatt
     5341 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     5401 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     5461 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     5521 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     5581 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
     5641 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     5701 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     5761 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     5821 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     5881 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     5941 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     6001 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     6061 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     6121 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     6181 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag
     6241 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     6301 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     6361 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     6421 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     6481 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     6541 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     6601 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     6661 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     6721 cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg
     6781 cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga
     6841 gcgaggaagc ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc
     6901 attaatgcag caagctcatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     6961 ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
     7021 caaaaagctc cccgtggcac gacaggtttc ccgactggaa agcgggcagt gagcgcaacg
     7081 caattaatgt gagttagctc actcattagg caccccaggc tttacacttt atgcttccgg
     7141 ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca cacaggaaac agctatgaca
     7201 tgattacgaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     7261 aactcatcaa tgtatcttat catgtctgga tcaactggat aactcaagct aaccaaaatc
     7321 atcccaaact tcccacccca taccctatta ccactgccaa ttacctgtgg tttcatttac
     7381 tctaaacctg tgattcctct gaattatttt cattttaaag aaattgtatt tgttaaatat
     7441 gtactacaaa cttagtagt
//
