LOCUS       sequence-315124-        7703 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of scFLARE1 component.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7703)
  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 7703)
  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: scFLARE1
FEATURES             Location/Qualifiers
     source          1..7703
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     553..572
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        553..571
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      585..594
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     591..650
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     CDS             1416..1925
                     /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             3612..3632
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQM"
     CDS             3645..3668
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             3669..4106
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae GAL4 (truncated)"
                     /product="DNA binding domain of the GAL4 transcriptional 
                     activator"
                     /label=GAL4 DNA binding domain
                     /translation="KLLSSIEQACDICRLKKLKCSKEKPKCAKCLKNNWECRYSPKTKR
                     SPLTRAHLTEVESRLERLEQLFLLIFPREDLDMILKMDSLQDIKALLTGLFVQDNVNKD
                     AVTDRLASVETDMPLTLRQHRISATSSSEESSNKGQRQLTVS"
     CDS             4107..4448
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae GAL4 (truncated)"
                     /product="activation domain of the GAL4 transcriptional 
                     activator"
                     /label=GAL4 activation domain
                     /translation="ANFNQSGNIADSSLSFTFTNSSNGPNLITTQTNSQALSQPIASSN
                     VHDNFMNNEITASKIDDGNNSKPLSPGWTDQTAYNAFGITTGMFNTTTMDDVYNYLFDD
                     EDTPPNPKKE"
     CDS             4449..4490
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(4524..4541)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4530..4737
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4766..4906
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4766..4895
                     /label=AAV2 ITR
     rep_origin      4981..5436
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5068..5087)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5278..5299
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5453..5472)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5572..5594
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5632..5650)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5718..5822
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5823..5891,5892..6683)
                     /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(6041..6060)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6854..7442
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7343..7362
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   7504..7633
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca ctgcttactg
      541 gcttatcgaa attaatacga ctcactatag ggagacccaa gcttgccacc atggagacag
      601 acacactcct gctatgggta ctgctgctct gggttccagg ttccactggt gacggtgccc
      661 agccggccct cgacttccag aaggcctcca gcatagtcta taagaaagag ggggaacagg
      721 tggagttctc cttcccactc gcctttacag ttgaaaagct gacgggcagt ggcgagctgt
      781 ggtggcaggc ggagagggct tcctcctcca agtcttggat cacctttgac ctgaagaaca
      841 aggaagtgtc tgtaaaacgg gttacccagg accctaagct ccagatgggc aagaagctcc
      901 cgctccacct caccctgccc caggccttgc ctcagtatgc tggctctgga aacctcaccc
      961 tggcccttga agcgaaaaca ggaaagttgc atcaggaagt gaacctggtg gtgatgagag
     1021 ccactcagct ccagaaaaat ttgacctgtg aggtgtgggg acccacctcc cctaagctga
     1081 tgctgagctt gaaactggag aacaaggagg caaaggtctc gaagcgggag aaggcggtgt
     1141 gggtgctgaa ccctgaggcg gggatgtggc agtgtctgct gagtgactcg ggacaggtcc
     1201 tgctggaatc caacatcaag gttctgccca catggtccac cccggtgcag ccaatggccc
     1261 tgattgtgct ggggggcgtc gccggcctcc tgcttttcat tgggctaggc atcttcttct
     1321 gtgtcaggtg ccggcaccga aggcgcaagg gctcgggctc gacctcgggc tcgggcagcg
     1381 gtggctctag aggttctggt ggcagctctg gaggtatgaa tggagctata ggaggtgacc
     1441 ttttgctcaa ttttcctgac atgtcggtcc tagagcgcca aagggctcac ctcaagtacc
     1501 tcaatcccac ctttgattct cctctcgccg gcttctttgc cgattcttca atgattaccg
     1561 gcggcgagat ggacagctat ctttcgactg ccggtttgaa tcttccgatg atgtacggtg
     1621 agacgacggt ggaaggtgat tcaagactct caatttcgcc ggaaacgacg cttgggactg
     1681 gaaatttcaa ggcagcgaag tttgatacag agactaagga ttgtaatgag gcggcgaaga
     1741 agatgacgat gaacagagat gacctagtag aagaaggaga agaagagaag tcgaaaataa
     1801 cagagcaaaa caatgggagc acaaaaagca tcaagaagat gaaacacaaa gccaagaaag
     1861 aagagaacaa tttctctaat gattcatcta aagtgacgaa ggaattggag aaaacggatt
     1921 atattcatgg tggctctggt agcggagaaa gcttgtttaa gggaccacgt gattacaacc
     1981 cgatatcgag caccatttgt catttgacga atgaatctga tgggcacaca acatcgttgt
     2041 atggtattgg atttggtccc ttcatcatta caaacaagca cttgtttaga agaaataatg
     2101 gaacactgtt ggtccaatca ctacatctcg aggaccaact gactgaagag cagatcgcag
     2161 aatttaaaga ggctttctcc ctatttgaca aggacgggga tgggacaata acaaccaagg
     2221 agctggggac ggtgatgcgg tctctggggc agaaccccac agaagcagag ctgcaggaca
     2281 tgatcaatga agtagatgcc gacggtgacg gcacaatcga cttccctgag ttcctgacaa
     2341 tgatggcaag aaaaatgaaa tacacaggcg gtgtgaagct gatcccctcc tggaccaccg
     2401 tgatcctggt gaagagcatg ctgcgcaagc gcagcttcgg caaccccttc ggtggcgaca
     2461 gtgaagaaga aattagagaa gcgttccgtg tgtttgataa ggatggcaat ggctacatca
     2521 gtgcagcaga gcttcgccac gtgatgacaa accttggaga gaagttaaca gatgaagagg
     2581 ttgatgaaat gatcagggaa gcagacatcg atggggatgg tcaggtaaac tacgaagagt
     2641 ttgtacaaat gatgacagcg aaggcggccg cgggtgtatt caaggtcaag aacaccacga
     2701 ctttgcaaca acacctcatt gatgggaggg acatgataat tattcgcatg cctaaggatt
     2761 tcccaccatt tcctcaaaag ctgaaattta gagagccaca aagggaagag cgcatatgtc
     2821 ttgtgacaac caacttccaa actaagagca tgtctagcat ggtgtcagac actagttgca
     2881 cattcccttc atctgatggc atattctgga agcattggat tcaaaccaag gatgggcagt
     2941 gtggcaatcc attagtatca actagagatg ggttcattgt tggtatacac tcagcatcga
     3001 atttcaccaa cacaaacaat tatttcacaa gcgtgccgaa aaacttcatg gaattgttga
     3061 caaatcagga ggcgcagcag tgggttagtg gttggcgatt aaatgctgac tcagtattgt
     3121 ggggtggcca taaagttttc atggtggagc tggccgagaa gctggccggc ctggacatca
     3181 acggtggagc tagcgagttt cgggcaacca cactggaacg gatcgagaaa agcttcgtga
     3241 ttactgatcc gagactgcct gacaacccaa tcatttttgt gagcgattcc ttcctgcagc
     3301 tgacagaata ttctcgggaa gagatcctgg ggcgcaattg ccgttttctg cagggacccg
     3361 agacagaccg tgccactgtt cggaaaatca gagatgctat tgacaaccag actgaagtga
     3421 ccgttcagct gatcaattat accaagagcg gcaagaagtt ctggaacgtg ttccacctgc
     3481 agccgatgcg cgattataag ggcgacgtcc agtacttcat tggcgtgcag ctggatggca
     3541 ccgaacgtct tcatggcgcc gctgagcgtg aggcggtctg cctggtcaaa aagacagcct
     3601 ttcagattgc tgagaacctg tacttccaga tgggcggagg atccgactac aaggatgacg
     3661 acgataagaa gctactgtct tctatcgaac aagcatgcga tatttgccga cttaaaaagc
     3721 tcaagtgctc caaagaaaaa ccgaagtgcg ccaagtgtct gaagaacaac tgggagtgtc
     3781 gctactctcc caaaaccaaa aggtctccgc tgactagggc acatctgaca gaagtggaat
     3841 caaggctaga aagactggaa cagctatttc tactgatttt tcctcgagaa gaccttgaca
     3901 tgattttgaa aatggattct ttacaggata taaaagcatt gttaacagga ttatttgtac
     3961 aagataatgt gaataaagat gccgtcacag atagattggc ttcagtggag actgatatgc
     4021 ctctaacatt gagacagcat agaataagtg cgacatcatc atcggaagag agtagtaaca
     4081 aaggtcaaag acagttgact gtatcggcca attttaatca aagtgggaat attgctgata
     4141 gctcattgtc cttcactttc actaacagta gcaacggtcc gaacctcata acaactcaaa
     4201 caaattctca agcgctttca caaccaattg cctcctctaa cgttcatgat aacttcatga
     4261 ataatgaaat cacggctagt aaaattgatg atggtaataa ttcaaaacca ctgtcacctg
     4321 gttggacgga ccaaactgcg tataacgcgt ttggaatcac tacagggatg tttaatacca
     4381 ctacaatgga tgatgtatat aactatctat tcgatgatga agatacccca ccaaacccaa
     4441 aaaaagaggg caagccgatc cctaaccctc tgctgggcct ggacagcact taagaattcc
     4501 tcgagctaga gctcgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt
     4561 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
     4621 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
     4681 ggtggggcag gacagcaagg gggaggattg ggaagacaat agcaggcatg ctggggaggt
     4741 aaccacgtgc ggaccgagcg gccgcaggaa cccctagtga tggagttggc cactccctct
     4801 ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt
     4861 gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg
     4921 tattttctcc ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag caaccatagt
     4981 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     5041 ctacacttgc cagcgcctta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     5101 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     5161 gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca cgtagtgggc
     5221 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     5281 gactcttgtt ccaaactgga acaacactca actctatctc gggctattct tttgatttat
     5341 aagggatttt gccgatttcg gtctattggt taaaaaatga gctgatttaa caaaaattta
     5401 acgcgaattt taacaaaata ttaacgttta caattttatg gtgcactctc agtacaatct
     5461 gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct
     5521 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     5581 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga
     5641 tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca
     5701 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     5761 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     5821 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     5881 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     5941 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     6001 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     6061 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     6121 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     6181 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     6241 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     6301 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     6361 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     6421 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     6481 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtggaa
     6541 gccgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     6601 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     6661 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     6721 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     6781 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     6841 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     6901 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     6961 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
     7021 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     7081 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     7141 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     7201 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     7261 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     7321 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     7381 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     7441 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     7501 tgtcctgcag gcagctgcgc gctcgctcgc tcactgaggc cgcccgggcg tcgggcgacc
     7561 tttggtcgcc cggcctcagt gagcgagcga gcgcgcagag agggagtggc caactccatc
     7621 actaggggtt cctgcggccg cacgcgttgt acatagtaat caattacggg gtcattagtt
     7681 catagcccat atatggagtt ccg
//
