LOCUS       sequence-318319-        7335 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Neuronal expression of scFLARE2 component.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7335)
  AUTHORS   Sanchez MI, Nguyen QA, Wang W, Soltesz I, Ting AY
  TITLE     Transcriptional readout of neuronal activity via an engineered 
            Ca(2+)-activated protease.
  JOURNAL   Proc Natl Acad Sci U S A. 2020 Dec 15. pii: 2006521117. doi: 
            10.1073/pnas.2006521117.
  PUBMED    33323488
REFERENCE   2  (bases 1 to 7335)
  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: "Proc Natl 
            Acad Sci U S A. 2020 Dec 15. pii: 2006521117. doi: 
            10.1073/pnas.2006521117."
COMMENT     Sequence Label: scFLARE2
FEATURES             Location/Qualifiers
     source          1..7335
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        181..628
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     CDS             811..837
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1063..1572
                     /codon_start=1
                     /product="N-terminal portion of the CIB1 transcription 
                     factor from Arabidopsis thaliana"
                     /label=CIBN
                     /note="binds to CRY2 (cryptochrome 2) that has absorbed 
                     blue light (Kennedy et al., 2010)"
                     /translation="MNGAIGGDLLLNFPDMSVLERQRAHLKYLNPTFDSPLAGFFADSS
                     MITGGEMDSYLSTAGLNLPMMYGETTVEGDSRLSISPETTLGTGNFKAAKFDTETKDCN
                     EAAKKMTMNRDDLVEEGEEEKSKITEQNNGSTKSIKKMKHKAKKEENNFSNDSSKVTKE
                     LEKTDYI"
     CDS             3259..3279
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQM"
     CDS             3292..3315
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             join(3316..3936,3937..4323)
                     /codon_start=1
                     /product="tetracycline-controlled transactivator, 
                     comprising a fusion of the tetracycline repressor TetR with
                     the C-terminal activation domain of herpes simplex virus 
                     VP16"
                     /label=tTA
                     /note="In the Tet-Off(R) system, tTA binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the absence of tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGSAYSRARTKNNYGSTI
                     EGLLDLPDDDAPEEAGLAAPRLSFLPAGHTRRLSTAPPTDVSLGDELHLDGEDVAMAHA
                     DALDDFDLDMLGDGDSPGPGFTPHDSAPYGALDMADFEFEQMFTDALGIDEYGG"
     primer_bind     complement(3389..3408)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(4357..4374)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4363..4570
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4599..4739
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4599..4728
                     /label=AAV2 ITR
     rep_origin      4813..5268
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4900..4919)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5110..5131
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5285..5304)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5404..5426
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5464..5482)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5550..5654
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5655..5723,5724..6515)
                     /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     complement(5873..5892)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6686..7274
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7175..7194
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgttgtaca gtgtctagac tgcagagggc cctgcgtatg
      181 agtgcaagtg ggttttagga ccaggatgag gcggggtggg ggtgcctacc tgacgaccga
      241 ccccgaccca ctggacaagc acccaacccc cattccccaa attgcgcatc ccctatcaga
      301 gagggggagg ggaaacagga tgcggcgagg cgcgtgcgca ctgccagctt cagcaccgcg
      361 gacagtgcct tcgcccccgc ctggcggcgc gcgccaccgc cgcctcagca ctgaaggcgc
      421 gctgacgtca ctcgccggtc ccccgcaaac tccccttccc ggccaccttg gtcgcgtccg
      481 cgccgccgcc ggcccagccg gaccgcacca cgcgaggcgc gagatagggg ggcacgggcg
      541 cgaccatctg cgctgcggcg ccggcgactc agcgctgcct cagtctgcgg tgggcagcgg
      601 aggagtcgtg tcgtgcctga gagcgcagtc gagaagaagc ttgccaccat gcacctgaga
      661 atccacgcga gacggagccc tcctcgccgg ccggcctgga cgcttgggat ctggttcctg
      721 ttctggggat gtatcgtcag ctctgtatgg agtcaattgt cttctaatgt agcttcctcc
      781 tcctccacct cttcctcgcc ggggtctcac tacccatacg atgtgccaga ttacgccaac
      841 cccacggagc cgggaatcag acgggttccg ggggcctcag aggtgatccg ggagtcgagc
      901 agcacaacag ggatggtcgt cggcattgtg gctgctgccg ccctctgcat cttgatcctc
      961 ctgtacgcca tgtacaagta caggaacagg gacgagggct cgggctcgac ctcgggctcg
     1021 ggcagcggtg gctctggagg ttctggtggc agctctggag gtatgaatgg agctatagga
     1081 ggtgaccttt tgctcaattt tcctgacatg tcggtcctag agcgccaaag ggctcacctc
     1141 aagtacctca atcccacctt tgattctcct ctcgccggct tctttgccga ttcttcaatg
     1201 attaccggcg gcgagatgga cagctatctt tcgactgccg gtttgaatct tccgatgatg
     1261 tacggtgaga cgacggtgga aggtgattca agactctcaa tttcgccgga aacgacgctt
     1321 gggactggaa atttcaaggc agcgaagttt gatacagaga ctaaggattg taatgaggcg
     1381 gcgaagaaga tgacgatgaa cagagatgac ctagtagaag aaggagaaga agagaagtcg
     1441 aaaataacag agcaaaacaa tgggagcaca aaaagcatca agaagatgaa acacaaagcc
     1501 aagaaagaag agaacaattt ctctaatgat tcatctaaag tgacgaagga attggagaaa
     1561 acggattata ttcatggtgg ctctggtagc ggagaaagct tgtttaaggg accacgtgat
     1621 tacaacccga tatcgagcac catttgtcat ttgacgaatg aatctgatgg gcacacaaca
     1681 tcgttgtatg gtattggatt tggtcccttc atcattacaa acaagcactt gtttagaaga
     1741 aataatggaa cactgttggt ccaatcacta catctcgagg accaactgac tgaagagcag
     1801 atcgcagaat ttaaagagct cttctcccta tttgacaagg acggggatgg gacaataaca
     1861 accaaggagc tggggacggt gatgcggtct ctggggcaga accccacaga agcagagctg
     1921 caagacatga tcaatgaagt agatgccgac ggtgacggca caatcgactt ccctgagttc
     1981 ctgaccatga tggctagaaa aatgaaggac acaggcggtg tgaagctgat cccctcctgg
     2041 accaccgtga tcctggtgaa gagcatgctg cgcaagcgca gcttcggcaa ccccttcggt
     2101 ggcgacagcg aagaggaaat ccgagaagca ttcggtgtgt ttgataagga tggcaatggc
     2161 tacatcagtg cagcagagct tcgccacgtg atgacaaacc ttggagagaa gttaacagat
     2221 gaagaggttg atgaaatgat cagggaagca gacatcgatg gggatggtca ggtaaactac
     2281 gaagagtttg tacaaatgat gacagcgaag gcggccgcgg gtgtattcaa ggtcaagaac
     2341 accacgactt tgcaacaaca cctcattgat gggagggaca tgataattat tcgcatgcct
     2401 aaggatttcc caccatttcc tcaaaagctg aaatttagag agccacaaag ggaagagcgc
     2461 atatgtcttg tgacaaccaa cttccaaact aagagcatgt ctagcatggt gtcagacact
     2521 agttgcacat tcccttcatc tgatggcata ttctggaagc attggattca aaccaaggat
     2581 gggcagtgtg gcaatccatt agtatcaact agagatgggt tcattgttgg tatacactca
     2641 gcatcgaatt tcaccaacac aaacaattat ttcacaagcg tgccgaaaaa cttcatggaa
     2701 ttgttgacaa atcaggaggc gcagcagtgg gttagtggtt ggcgattaaa tgctgactca
     2761 gtattgtggg gtggccataa agttttcatg gtggagctgg ccgagaagct ggccggcctg
     2821 gacatcaacg gtggagctag cgagtttcgg gcaaccacac tggaacggat cgagaaaagc
     2881 ttcgtgatta ctgatccgag actgcctgac aacccaatca tttttgtgag cgattccttc
     2941 ctgcagctga cagaatattc tcgggaagag atcctggggc gcaattgccg ttttctgcag
     3001 ggacccgaga cagaccgtgc cactgttcgg aaaatcagag atgctattga caaccagact
     3061 gaagtgaccg ttcagctgat caattatacc aagagcggca agaagttctg gaacgtgttc
     3121 cacctgcagc cgatgcgcga ttataagggc gacgtccagt acttcattgg cgtgcagctg
     3181 gatggcaccg aacgtcttca tggcgccgct gagcgtgagg cggtctgcct ggtcaaaaag
     3241 acagcctttc agattgctga gaacctgtac ttccagatgg gcggaggatc cgactacaag
     3301 gatgacgacg ataagatgtc tagattagat aaaagtaaag tgattaacag cgcattagag
     3361 ctgcttaatg aggtcggaat cgaaggttta acaacccgta aactcgccca gaagctaggt
     3421 gtagagcagc ctacattgta ttggcatgta aaaaataagc gggctttgct cgacgcctta
     3481 gccattgaga tgttagatag gcaccatact cacttttgcc ctttagaagg ggaaagctgg
     3541 caagattttt tacgtaataa cgctaaaagt tttagatgtg ctttactaag tcatcgcgat
     3601 ggagcaaaag tacatttagg tacacggcct acagaaaaac agtatgaaac tctcgaaaat
     3661 caattagcct ttttatgcca acaaggtttt tcactagaga atgcattata tgcactcagc
     3721 gctgtggggc attttacttt aggttgcgta ttggaagatc aagagcatca agtcgctaaa
     3781 gaagaaaggg aaacacctac tactgatagt atgccgccat tattacgaca agctatcgaa
     3841 ttatttgatc accaaggtgc agagccagcc ttcttattcg gccttgaatt gatcatatgc
     3901 ggattagaaa aacaacttaa atgtgaaagt gggtccgcgt acagccgcgc gcgtacgaaa
     3961 aacaattacg ggtctaccat cgagggcctg ctcgatctcc cggacgacga cgcccccgaa
     4021 gaggcggggc tggcggctcc gcgcctgtcc tttctccccg cgggacacac gcgcagactg
     4081 tcgacggccc ccccgaccga tgtcagcctg ggggacgagc tccacttaga cggcgaggac
     4141 gtggcgatgg cgcatgccga cgcgctagac gatttcgatc tggacatgtt gggggacggg
     4201 gattccccgg gtccgggatt taccccccac gactccgccc cctacggcgc tctggatatg
     4261 gccgacttcg agtttgagca gatgtttacc gatgcccttg gaattgacga gtacggtggg
     4321 taataggaat tcctcgagct agagctcgct gatcagcctc gactgtgcct tctagttgcc
     4381 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
     4441 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
     4501 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc
     4561 atgctgggga ggtaaccacg tgcggaccga gcggccgcag gaacccctag tgatggagtt
     4621 ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg
     4681 acgcccgggc tttgcccggg cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg
     4741 gcgcctgatg cggtattttc tccttacgca tctgtgcggt attcacaccg catacgtcaa
     4801 agcaaccata gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     4861 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     4921 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     4981 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgatttg ggtgatggtt
     5041 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     5101 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcgggctatt
     5161 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     5221 aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt tacaatttta tggtgcactc
     5281 tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg
     5341 ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg
     5401 tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa
     5461 agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga
     5521 cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa
     5581 tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt
     5641 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     5701 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     5761 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     5821 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     5881 gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
     5941 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     6001 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     6061 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     6121 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     6181 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
     6241 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     6301 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     6361 gtgagcgtgg aagccgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     6421 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     6481 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata
     6541 tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt
     6601 ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc
     6661 ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct
     6721 tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa
     6781 ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag
     6841 tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc
     6901 tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg
     6961 actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca
     7021 cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat
     7081 gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg
     7141 tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc
     7201 ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc
     7261 ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc
     7321 cttttgctca catgt
//
