LOCUS       sequence-267519-        6364 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Neuronal expression of uFLARE TF component.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6364)
  AUTHORS   Sanchez MI, Ting AY
  TITLE     Directed evolution improves the catalytic efficiency of TEV 
            protease.
  JOURNAL   Nat Methods. 2019 Dec 9. pii: 10.1038/s41592-019-0665-7. doi: 
            10.1038/s41592-019-0665-7.
  PUBMED    31819267
REFERENCE   2  (bases 1 to 6364)
  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 
            Methods. 2019 Dec 9. pii: 10.1038/s41592-019-0665-7. doi: 
            10.1038/s41592-019-0665-7."
COMMENT     Sequence Label: 
            Syn:NRX-TM-Nav1.6-CIBN-2xMKII-hLOV-TEcs(ENLYFQ/M)-tTA
FEATURES             Location/Qualifiers
     source          1..6364
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             628..654
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             880..1389
                     /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             2104..2124
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQM"
     CDS             2137..2160
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             join(2161..2781,2782..3168)
                     /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(2234..2253)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     primer_bind     complement(3202..3219)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3208..3415
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3444..3584
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3444..3573
                     /label=AAV2 ITR
     rep_origin      3659..4114
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3746..3765)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3956..3977
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4131..4150)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4250..4272
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4310..4328)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4396..4500
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4501..4569,4570..5361)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEVDKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(4719..4738)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5532..6120
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6021..6040
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   6182..6311
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        join(6362..6364,1..445)
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
ORIGIN
        1 gcaagtgggt tttaggacca ggatgaggcg gggtgggggt gcctacctga cgaccgaccc
       61 cgacccactg gacaagcacc caacccccat tccccaaatt gcgcatcccc tatcagagag
      121 ggggagggga aacaggatgc ggcgaggcgc gtgcgcactg ccagcttcag caccgcggac
      181 agtgccttcg cccccgcctg gcggcgcgcg ccaccgccgc ctcagcactg aaggcgcgct
      241 gacgtcactc gccggtcccc cgcaaactcc ccttcccggc caccttggtc gcgtccgcgc
      301 cgccgccggc ccagccggac cgcaccacgc gaggcgcgag ataggggggc acgggcgcga
      361 ccatctgcgc tgcggcgccg gcgactcagc gctgcctcag tctgcggtgg gcagcggagg
      421 agtcgtgtcg tgcctgagag cgcagtcgag aagaagcttg ccaccatgca cctgagaatc
      481 cacgcgagac ggagccctcc tcgccggccg gcctggacgc ttgggatctg gttcctgttc
      541 tggggatgta tcgtcagctc tgtatggagt caattgtctt ctaatgtagc ttcctcctcc
      601 tccacctctt cctcgccggg gtctcactac ccatacgatg tgccagatta cgccaacccc
      661 acggagccgg gaatcagacg ggttccgggg gcctcagagg tgatccggga gtcgagcagc
      721 acaacaggga tggtcgtcgg cattgtggct gctgccgccc tctgcatctt gatcctcctg
      781 tacgccatgt acaagtacag gaacagggac gagggctcgg gctcgacctc gggctcgggc
      841 agcggtggct ctggaggttc tggtggcagc tctggaggta tgaatggagc tataggaggt
      901 gaccttttgc tcaattttcc tgacatgtcg gtcctagagc gccaaagggc tcacctcaag
      961 tacctcaatc ccacctttga ttctcctctc gccggcttct ttgccgattc ttcaatgatt
     1021 accggcggcg agatggacag ctatctttcg actgccggtt tgaatcttcc gatgatgtac
     1081 ggtgagacga cggtggaagg tgattcaaga ctctcaattt cgccggaaac gacgcttggg
     1141 actggaaatt tcaaggcagc gaagtttgat acagagacta aggattgtaa tgaggcggcg
     1201 aagaagatga cgatgaacag agatgaccta gtagaagaag gagaagaaga gaagtcgaaa
     1261 ataacagagc aaaacaatgg gagcacaaaa agcatcaaga agatgaaaca caaagccaag
     1321 aaagaagaga acaatttctc taatgattca tctaaagtga cgaaggaatt ggagaaaacg
     1381 gattatattc atagcggtag tggcaccgtg cgcgtgccga ttgcggtggg cgaaagcgat
     1441 tttgaaaacc tgaacaccga agatgtgagc agcgaaagcg atccgggtgg ctctggtagc
     1501 ttcaacgctc gcaggaagct ggctggtgct atcctgttca ccatgctcgc cacaagaaac
     1561 ttcagcggca gctttaatgc gcggcgcaag ctggccggag ccatactttt cacaatgttg
     1621 gctaccagaa atttttcgga gctggccgag aagctggccg gcctggacat caacggtgga
     1681 gctagcgagt ttcgggcaac cacactggaa cggatcgaga aaagcttcgt gattactgat
     1741 ccgagactgc ctgacaaccc aatcattttt gtgagcgatt ccttcctgca gctgacagaa
     1801 tattctcggg aagagatcct ggggcgcaat tgccgttttc tgcagggacc cgagacagac
     1861 cgtgccactg ttcggaaaat cagagatgct attgacaacc agactgaagt gaccgttcag
     1921 ctgatcaatt ataccaagag cggcaagaag ttctggaacg tgttccacct gcagccgatg
     1981 cgcgattata agggcgacgt ccagtacttc attggcgtgc agctggatgg caccgaacgt
     2041 cttcatggcg ccgctgagcg tgaggcggtc tgcctggtca aaaagacagc ctttcagatt
     2101 gctgagaacc tgtacttcca gatgggcgga ggatccgact acaaggatga cgacgataag
     2161 atgtctagat tagataaaag taaagtgatt aacagcgcat tagagctgct taatgaggtc
     2221 ggaatcgaag gtttaacaac ccgtaaactc gcccagaagc taggtgtaga gcagcctaca
     2281 ttgtattggc atgtaaaaaa taagcgggct ttgctcgacg ccttagccat tgagatgtta
     2341 gataggcacc atactcactt ttgcccttta gaaggggaaa gctggcaaga ttttttacgt
     2401 aataacgcta aaagttttag atgtgcttta ctaagtcatc gcgatggagc aaaagtacat
     2461 ttaggtacac ggcctacaga aaaacagtat gaaactctcg aaaatcaatt agccttttta
     2521 tgccaacaag gtttttcact agagaatgca ttatatgcac tcagcgctgt ggggcatttt
     2581 actttaggtt gcgtattgga agatcaagag catcaagtcg ctaaagaaga aagggaaaca
     2641 cctactactg atagtatgcc gccattatta cgacaagcta tcgaattatt tgatcaccaa
     2701 ggtgcagagc cagccttctt attcggcctt gaattgatca tatgcggatt agaaaaacaa
     2761 cttaaatgtg aaagtgggtc cgcgtacagc cgcgcgcgta cgaaaaacaa ttacgggtct
     2821 accatcgagg gcctgctcga tctcccggac gacgacgccc ccgaagaggc ggggctggcg
     2881 gctccgcgcc tgtcctttct ccccgcggga cacacgcgca gactgtcgac ggcccccccg
     2941 accgatgtca gcctggggga cgagctccac ttagacggcg aggacgtggc gatggcgcat
     3001 gccgacgcgc tagacgattt cgatctggac atgttggggg acggggattc cccgggtccg
     3061 ggatttaccc cccacgactc cgccccctac ggcgctctgg atatggccga cttcgagttt
     3121 gagcagatgt ttaccgatgc ccttggaatt gacgagtacg gtgggtaata ggaattcctc
     3181 gagctagagc tcgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt
     3241 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     3301 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     3361 tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct ggggaggtaa
     3421 ccacgtgcgg accgagcggc cgcaggaacc cctagtgatg gagttggcca ctccctctct
     3481 gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc gcccgacgcc cgggctttgc
     3541 ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg caggggcgcc tgatgcggta
     3601 ttttctcctt acgcatctgt gcggtatttc acaccgcata cgtcaaagca accatagtac
     3661 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct
     3721 acacttgcca gcgccttagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg
     3781 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt
     3841 gctttacggc acctcgaccc caaaaaactt gatttgggtg atggttcacg tagtgggcca
     3901 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga
     3961 ctcttgttcc aaactggaac aacactcaac tctatctcgg gctattcttt tgatttataa
     4021 gggattttgc cgatttcggt ctattggtta aaaaatgagc tgatttaaca aaaatttaac
     4081 gcgaatttta acaaaatatt aacgtttaca attttatggt gcactctcag tacaatctgc
     4141 tctgatgccg catagttaag ccagccccga cacccgccaa cacccgctga cgcgccctga
     4201 cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc cgggagctgc
     4261 atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg cctcgtgata
     4321 cgcctatttt tataggttaa tgtcatgata ataatggttt cttagacgtc aggtggcact
     4381 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     4441 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     4501 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     4561 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
     4621 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
     4681 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
     4741 cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
     4801 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
     4861 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     4921 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     4981 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     5041 cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     5101 tcccggcaac aattaataga ctggatggag gtggataaag ttgcaggacc acttctgcgc
     5161 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtggaagc
     5221 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     5281 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     5341 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     5401 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     5461 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     5521 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     5581 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     5641 gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta
     5701 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     5761 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     5821 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     5881 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     5941 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     6001 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     6061 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     6121 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     6181 tcctgcaggc agctgcgcgc tcgctcgctc actgaggccg cccgggcgtc gggcgacctt
     6241 tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca actccatcac
     6301 taggggttcc tgcggccgca cgcgttgtac agtgtctaga ctgcagaggg ccctgcgtat
     6361 gagt
//
