LOCUS       sequence-320945-        6659 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  FLiCRE TF 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-hLOV1-TEVcs(ENLYFQ/M)- Gal4
FEATURES             Location/Qualifiers
     source          1..6659
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        71..374
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        375..578
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     529..549
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     623..642
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        623..641
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      655..664
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     661..720
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     CDS             745..771
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             772..798
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1546..2055
                     /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             2677..2697
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQM"
     CDS             2710..2733
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2734..3171
                     /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             3172..3516
                     /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(3550..3567)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3556..3763
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3792..3932
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3792..3921
                     /label=AAV2 ITR
     rep_origin      4007..4462
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4094..4113)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4304..4325
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4479..4498)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4598..4620
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4658..4676)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4744..4848
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4849..4917,4918..5709)
                     /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(5067..5086)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5880..6468
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6369..6388
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   6530..6659
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
ORIGIN
        1 gcggccgcac gcgttgtaca tagtaatcaa ttacggggtc attagttcat agcccatata
       61 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      121 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      181 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      241 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      301 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      361 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      421 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      481 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      541 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      601 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gcttgccacc
      661 atggagacag acacactcct gctatgggta ctgctgctct gggttccagg ttccactggt
      721 gacggggccc agccggccag atcttaccca tacgatgtgc cagattacgc ctatccctat
      781 gacgtccctg attacgctct cgacttccag aaggcctcca gcatagtcta taagaaagag
      841 ggggaacagg tggagttctc cttcccactc gcctttacag ttgaaaagct gacgggcagt
      901 ggcgagctgt ggtggcaggc ggagagggct tcctcctcca agtcttggat cacctttgac
      961 ctgaagaaca aggaagtgtc tgtaaaacgg gttacccagg accctaagct ccagatgggc
     1021 aagaagctcc cgctccacct caccctgccc caggccttgc ctcagtatgc tggctctgga
     1081 aacctcaccc tggcccttga agcgaaaaca ggaaagttgc atcaggaagt gaacctggtg
     1141 gtgatgagag ccactcagct ccagaaaaat ttgacctgtg aggtgtgggg acccacctcc
     1201 cctaagctga tgctgagctt gaaactggag aacaaggagg caaaggtctc gaagcgggag
     1261 aaggcggtgt gggtgctgaa ccctgaggcg gggatgtggc agtgtctgct gagtgactcg
     1321 ggacaggtcc tgctggaatc caacatcaag gttctgccca catggtccac cccggtgcag
     1381 ccaatggccc tgattgtgct ggggggcgtc gccggcctcc tgcttttcat tgggctaggc
     1441 atcttcttct gtgtcaggtg ccggcaccga aggcgcaagg gctcgggctc gacctcgggc
     1501 tcgggcagcg gtggctctag aggttctggt ggcagctctg gaggtatgaa tggagctata
     1561 ggaggtgacc ttttgctcaa ttttcctgac atgtcggtcc tagagcgcca aagggctcac
     1621 ctcaagtacc tcaatcccac ctttgattct cctctcgccg gcttctttgc cgattcttca
     1681 atgattaccg gcggcgagat ggacagctat ctttcgactg ccggtttgaa tcttccgatg
     1741 atgtacggtg agacgacggt ggaaggtgat tcaagactct caatttcgcc ggaaacgacg
     1801 cttgggactg gaaatttcaa ggcagcgaag tttgatacag agactaagga ttgtaatgag
     1861 gcggcgaaga agatgacgat gaacagagat gacctagtag aagaaggaga agaagagaag
     1921 tcgaaaataa cagagcaaaa caatgggagc acaaaaagca tcaagaagat gaaacacaaa
     1981 gccaagaaag aagagaacaa tttctctaat gattcatcta aagtgacgaa ggaattggag
     2041 aaaacggatt atattcatgg tggctctggt agcttcaacg ctcgcaggaa gctggctggt
     2101 gctatcctgt tcaccatgct cgccacaaga aacttcagcg gcagctttaa tgcgcggcgc
     2161 aagttggccg gagccatact tttcacaatg ttggctacca gaaatttttc ggagctggcc
     2221 gagaagctgg ccggcctgga catcaacggt ggagctagcg agtttcgggc aaccacactg
     2281 gaacggatcg agaaaagctt cgtgattact gatccgagac tgcctgacaa cccaatcatt
     2341 tttgtgagcg attccttcct gcagctgaca gaatattctc gggaagagat cctggggcgc
     2401 aattgccgtt ttctgcaggg acccgagaca gaccgtgcca ctgttcggaa aatcagagat
     2461 gctattgaca accagactga agtgaccgtt cagctgatca attataccaa gagcggcaag
     2521 aagttctgga acgtgttcca cctgcagccg atgcgcgatt ataagggcga cgtccagtac
     2581 ttcattggcg tgcagctgga tggcaccgaa cgtcttcatg gcgccgctga gcgtgaggcg
     2641 gtctgcctgg tcaaaaagac agcctttcag attgctgaga acctgtactt ccagatgggc
     2701 ggaggatccg actacaagga tgacgacgat aagaagctac tgtcttctat cgaacaagca
     2761 tgcgatattt gccgacttaa aaagctcaag tgctccaaag aaaaaccgaa gtgcgccaag
     2821 tgtctgaaga acaactggga gtgtcgctac tctcccaaaa ccaaaaggtc tccgctgact
     2881 agggcacatc tgacagaagt ggaatcaagg ctagaaagac tggaacagct atttctactg
     2941 atttttcctc gagaagacct tgacatgatt ttgaaaatgg attctttaca ggatataaaa
     3001 gcattgttaa caggattatt tgtacaagat aatgtgaata aagatgccgt cacagataga
     3061 ttggcttcag tggagactga tatgcctcta acattgagac agcatagaat aagtgcgaca
     3121 tcatcatcgg aagagagtag taacaaaggt caaagacagt tgactgtatc ggccaatttt
     3181 aatcaaagtg ggaatattgc tgatagctca ttgtccttca ctttcactaa cagtagcaac
     3241 ggtccgaacc tcataacaac tcaaacaaat tctcaagcgc tttcacaacc aattgcctcc
     3301 tctaacgttc atgataactt catgaataat gaaatcacgg ctagtaaaat tgatgatggt
     3361 aataattcaa aaccactgtc acctggttgg acggaccaaa ctgcgtataa cgcgtttgga
     3421 atcactacag ggatgtttaa taccactaca atggatgatg tatataacta tctattcgat
     3481 gatgaagata ccccaccaaa cccaaaaaaa gagtagtaag aattcctcga gctagagctc
     3541 gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg
     3601 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa
     3661 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca
     3721 gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggaggtaacc acgtgcggac
     3781 cgagcggccg caggaacccc tagtgatgga gttggccact ccctctctgc gcgctcgctc
     3841 gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc
     3901 agtgagcgag cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt ttctccttac
     3961 gcatctgtgc ggtatttcac accgcatacg tcaaagcaac catagtacgc gccctgtagc
     4021 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     4081 gccttagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     4141 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     4201 ctcgacccca aaaaacttga tttgggtgat ggttcacgta gtgggccatc gccctgatag
     4261 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     4321 actggaacaa cactcaactc tatctcgggc tattcttttg atttataagg gattttgccg
     4381 atttcggtct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac
     4441 aaaatattaa cgtttacaat tttatggtgc actctcagta caatctgctc tgatgccgca
     4501 tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg
     4561 ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg
     4621 ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta
     4681 taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat
     4741 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     4801 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
     4861 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
     4921 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
     4981 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
     5041 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
     5101 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
     5161 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
     5221 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     5281 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     5341 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg
     5401 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     5461 ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     5521 gctggctggt ttattgctga taaatctgga gccggtgagc gtggaagccg cggtatcatt
     5581 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     5641 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     5701 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
     5761 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     5821 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     5881 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     5941 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     6001 agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc
     6061 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     6121 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     6181 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     6241 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     6301 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     6361 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     6421 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     6481 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgtc ctgcaggcag
     6541 ctgcgcgctc gctcgctcac tgaggccgcc cgggcgtcgg gcgacctttg gtcgcccggc
     6601 ctcagtgagc gagcgagcgc gcagagaggg agtggccaac tccatcacta ggggttcct
//
