LOCUS       sequence-321437-        6659 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  FLiCRE TF with fast kinetics for HEK293T expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6659)
  AUTHORS   Kim CK, Sanchez MI, Hoerbelt P, Fenno LE, Malenka RC, Deisseroth K, 
            Ting AY
  TITLE     A Molecular Calcium Integrator Reveals a Striatal Cell Type Driving 
            Aversion.
  JOURNAL   Cell. 2020 Dec 23;183(7):2003-2019.e16. doi: 
            10.1016/j.cell.2020.11.015. Epub 2020 Dec 11.
  PUBMED    33308478
REFERENCE   2  (bases 1 to 6659)
  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.2020.11.015"; journalName: "Cell"; date: 
            "2020-12-23- 23"; volume: "183"; issue: "7"; pages: "2003-2019.e16"
COMMENT     Sequence Label: CD4-MKII-f-hLOV1-TEVcs(ENLYFQ/M)- Gal4
FEATURES             Location/Qualifiers
     source          1..6659
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     enhancer        201..504
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        505..708
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     659..679
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     753..772
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        753..771
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      785..794
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     791..850
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     CDS             875..901
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             902..928
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1676..2185
                     /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             2807..2827
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQM"
     CDS             2840..2863
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2864..3301
                     /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             3302..3646
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae GAL4 (truncated)"
                     /product="activation domain of the GAL4 transcriptional 
                     activator"
                     /label=GAL4 activation domain
                     /translation="ANFNQSGNIADSSLSFTFTNSSNGPNLITTQTNSQALSQPIASSN
                     VHDNFMNNEITASKIDDGNNSKPLSPGWTDQTAYNAFGITTGMFNTTTMDDVYNYLFDD
                     EDTPPNPKKE"
     primer_bind     complement(3680..3697)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3686..3893
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3922..4062
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3922..4051
                     /label=AAV2 ITR
     rep_origin      4137..4592
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4224..4243)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4434..4455
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4609..4628)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4728..4750
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4788..4806)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4874..4978
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4979..5047,5048..5839)
                     /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(5197..5216)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6010..6598
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6499..6518
                     /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 tagtaatcaa ttacggggtc attagttcat
      181 agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg
      241 cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata
      301 gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta
      361 catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc
      421 gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca gtacatctac
      481 gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga
      541 tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg
      601 ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg
      661 caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact
      721 agagaaccca ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa
      781 gcttgccacc atggagacag acacactcct gctatgggta ctgctgctct gggttccagg
      841 ttccactggt gacggggccc agccggccag atcttaccca tacgatgtgc cagattacgc
      901 ctatccctat gacgtccctg attacgctct cgacttccag aaggcctcca gcatagtcta
      961 taagaaagag ggggaacagg tggagttctc cttcccactc gcctttacag ttgaaaagct
     1021 gacgggcagt ggcgagctgt ggtggcaggc ggagagggct tcctcctcca agtcttggat
     1081 cacctttgac ctgaagaaca aggaagtgtc tgtaaaacgg gttacccagg accctaagct
     1141 ccagatgggc aagaagctcc cgctccacct caccctgccc caggccttgc ctcagtatgc
     1201 tggctctgga aacctcaccc tggcccttga agcgaaaaca ggaaagttgc atcaggaagt
     1261 gaacctggtg gtgatgagag ccactcagct ccagaaaaat ttgacctgtg aggtgtgggg
     1321 acccacctcc cctaagctga tgctgagctt gaaactggag aacaaggagg caaaggtctc
     1381 gaagcgggag aaggcggtgt gggtgctgaa ccctgaggcg gggatgtggc agtgtctgct
     1441 gagtgactcg ggacaggtcc tgctggaatc caacatcaag gttctgccca catggtccac
     1501 cccggtgcag ccaatggccc tgattgtgct ggggggcgtc gccggcctcc tgcttttcat
     1561 tgggctaggc atcttcttct gtgtcaggtg ccggcaccga aggcgcaagg gctcgggctc
     1621 gacctcgggc tcgggcagcg gtggctctag aggttctggt ggcagctctg gaggtatgaa
     1681 tggagctata ggaggtgacc ttttgctcaa ttttcctgac atgtcggtcc tagagcgcca
     1741 aagggctcac ctcaagtacc tcaatcccac ctttgattct cctctcgccg gcttctttgc
     1801 cgattcttca atgattaccg gcggcgagat ggacagctat ctttcgactg ccggtttgaa
     1861 tcttccgatg atgtacggtg agacgacggt ggaaggtgat tcaagactct caatttcgcc
     1921 ggaaacgacg cttgggactg gaaatttcaa ggcagcgaag tttgatacag agactaagga
     1981 ttgtaatgag gcggcgaaga agatgacgat gaacagagat gacctagtag aagaaggaga
     2041 agaagagaag tcgaaaataa cagagcaaaa caatgggagc acaaaaagca tcaagaagat
     2101 gaaacacaaa gccaagaaag aagagaacaa tttctctaat gattcatcta aagtgacgaa
     2161 ggaattggag aaaacggatt atattcatgg tggctctggt agcttcaacg ctcgcaggaa
     2221 gctggctggt gctatcctgt tcaccatgct cgccacaaga aacttcagcg gcagctttaa
     2281 tgcgcggcgc aagttggccg gagccatact tttcacaatg ttggctacca gaaatttttc
     2341 ggagctggcc gagaagctgg ccggcctgga catcaacggt ggagctagcg agtttcgggc
     2401 aaccacactg gaacggatcg agaaaagctt cgtgattact gatccgagac tgcctgacaa
     2461 cccagtgatt tttgtgagcg attccttcct gcagctgaca gaatattctc gggaagagat
     2521 cctggggcgc aattgccgtt ttctgcaggg acccgagaca gaccgtgcca ctgttcggaa
     2581 aatcagagat gctattgaca accagactga agtgaccgtt cagctgatca attataccaa
     2641 gagcggcaag aagttctgga acgtgttcca cctgcagccg atgcgcgatt ataagggcga
     2701 cgtccagtac ttcattggcg tgcagctgga tggcaccgaa cgtcttcatg gcgccgctga
     2761 gcgtgaggcg gtctgcctgg tcaaaaagac agcctttcag attgctgaga acctgtactt
     2821 ccagatgggc ggaggatccg actacaagga tgacgacgat aagaagctac tgtcttctat
     2881 cgaacaagca tgcgatattt gccgacttaa aaagctcaag tgctccaaag aaaaaccgaa
     2941 gtgcgccaag tgtctgaaga acaactggga gtgtcgctac tctcccaaaa ccaaaaggtc
     3001 tccgctgact agggcacatc tgacagaagt ggaatcaagg ctagaaagac tggaacagct
     3061 atttctactg atttttcctc gagaagacct tgacatgatt ttgaaaatgg attctttaca
     3121 ggatataaaa gcattgttaa caggattatt tgtacaagat aatgtgaata aagatgccgt
     3181 cacagataga ttggcttcag tggagactga tatgcctcta acattgagac agcatagaat
     3241 aagtgcgaca tcatcatcgg aagagagtag taacaaaggt caaagacagt tgactgtatc
     3301 ggccaatttt aatcaaagtg ggaatattgc tgatagctca ttgtccttca ctttcactaa
     3361 cagtagcaac ggtccgaacc tcataacaac tcaaacaaat tctcaagcgc tttcacaacc
     3421 aattgcctcc tctaacgttc atgataactt catgaataat gaaatcacgg ctagtaaaat
     3481 tgatgatggt aataattcaa aaccactgtc acctggttgg acggaccaaa ctgcgtataa
     3541 cgcgtttgga atcactacag ggatgtttaa taccactaca atggatgatg tatataacta
     3601 tctattcgat gatgaagata ccccaccaaa cccaaaaaaa gagtagtaag aattcctcga
     3661 gctagagctc gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc
     3721 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa
     3781 aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg
     3841 gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggaggtaacc
     3901 acgtgcggac cgagcggccg caggaacccc tagtgatgga gttggccact ccctctctgc
     3961 gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc
     4021 gggcggcctc agtgagcgag cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt
     4081 ttctccttac gcatctgtgc ggtatttcac accgcatacg tcaaagcaac catagtacgc
     4141 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     4201 acttgccagc gccttagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     4261 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     4321 tttacggcac ctcgacccca aaaaacttga tttgggtgat ggttcacgta gtgggccatc
     4381 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     4441 cttgttccaa actggaacaa cactcaactc tatctcgggc tattcttttg atttataagg
     4501 gattttgccg atttcggtct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     4561 gaattttaac aaaatattaa cgtttacaat tttatggtgc actctcagta caatctgctc
     4621 tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg
     4681 ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat
     4741 gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg
     4801 cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt
     4861 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     4921 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat
     4981 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt
     5041 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg
     5101 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
     5161 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
     5221 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
     5281 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
     5341 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg
     5401 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
     5461 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
     5521 tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc
     5581 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
     5641 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtggaagccg
     5701 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
     5761 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
     5821 actgattaag cattggtaac tgtcagacca agtttactca tatatacttt agattgattt
     5881 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     5941 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     6001 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     6061 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     6121 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg
     6181 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     6241 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     6301 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     6361 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     6421 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     6481 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     6541 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     6601 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgt
//
