LOCUS       sequence-481815-        7360 bp    DNA     circular SYN 08-AUG-2025
DEFINITION  Blue-light-controlled nuclear translocation of split tTA 
            transactivator from the cytoplasm. Split version..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7360)
  TITLE     Photoswitchable intein for light control of covalent protein binding
            and cleavage
REFERENCE   2  (bases 1 to 7360)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 8, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     Sequence Label: Split PS Intein tTA
FEATURES             Location/Qualifiers
     source          1..7360
                     /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"
     CDS             825..878
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             882..1817
                     /codon_start=1
                     /product="mutated ligand-binding domain of the human 
                     estrogen receptor (Feil et al., 1997)"
                     /label=ERT2
                     /translation="AGDMRAANLWPSPLMIKRSKKNSLALSLTADQMVSALLDAEPPIL
                     YSEYDPTRPFSEASMMGLLTNLADRELVHMINWAKRVPGFVDLTLHDQVHLLECAWLEI
                     LMIGLVWRSMEHPVKLLFAPNLLLDRNQGKCVEGMVEIFDMLLATSSRFRMMNLQGEEF
                     VCLKSIILLNSGVYTFLSSTLKSLEEKDHIHRVLDKITDTLIHLMAKAGLTLQQQHQRL
                     AQLLLILSHIRHMSNKGMEHLYSMKCKNVVPLYDLLLEAADAHRLHAPTSRGGASVEET
                     DQSHLATAGSTSSHSLQKYYITGEAEGFPAT"
     CDS             2424..2873
                     /codon_start=1
                     /product="""positive Magnet"" variant of an N-terminally 
                     truncated Vivid (VVD) blue light photoreceptor from 
                     Neurospora crassa (Kawano et al., 2015)"
                     /label=pMag
                     /note="rapidly heterodimerizes with nMagHigh1 upon blue 
                     light irradiation to form a complex with slow switch-off 
                     kinetics"
                     /translation="HTLYAPGGYDIMGYLIQIMNRPNPQVELGPVDTSCALILCDLKQK
                     DTPIVYASEAFLYMTGYSNAEVLGRNCRFLQSPDGMVKPKSTRKYVDSNTINTMRKAID
                     RNAEVQVEVVNFKKNGQRFVNFLTMIPVRDETGEYRYSMGFQCETE"
     CDS             join(2961..3578,3579..3965)
                     /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="SRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHVK
                     NKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTRP
                     TEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTTD
                     SMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGSAYSRARTKNNYGSTIE
                     GLLDLPDDDAPEEAGLAAPRLSFLPAGHTRRLSTAPPTDVSLGDELHLDGEDVAMAHAD
                     ALDDFDLDMLGDGDSPGPGFTPHDSAPYGALDMADFEFEQMFTDALGIDEYGG"
     primer_bind     complement(3031..3050)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     polyA_signal    4088..4209
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4125..4144)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4179..4198
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(4216..4671)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4353..4374)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4565..4584
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4698..4802
                     /gene="bla"
                     /label=AmpR promoter
     promoter        4804..5161
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        4804..4995
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(4827..4847)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      5012..5147
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             5196..5990
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             5196..5990
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5250..5269)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5860..5879
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(6178..6197)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6222..6269
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6598..7186
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7087..7106
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
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 gcagagctgg tttagtgaac cgtcagatcc gctagcacca
      541 tgagcggcta ttgtctggat ctgaaaaccc aggttcagac accgcagggt atgaaagaaa
      601 tttcaaatat tcaggtgggt gatctggttc tgagcaatac tggttataat gaagtgctga
      661 atgtgtttcc gaaaagcaaa aaaaaaagct ataaaatcac cctggaagat ggcaaagaaa
      721 tcatttgtag cgaagaacac ctgtttccga cccagactgg tgaaatgaat attagcggtg
      781 gtctgaaaga aggtatgtgc ctgtatgtta aatccggaag cggcgagggc aggggcagcc
      841 tgctgacctg cggcgacgtg gaggagaacc ccggccccgg cgctggagac atgagagctg
      901 ccaacctttg gccaagcccg ctcatgatca aacgctctaa gaagaacagc ctggccttgt
      961 ccctgacggc cgaccagatg gtcagtgcct tgttggatgc tgagcccccc atactctatt
     1021 ccgagtatga tcctaccaga cccttcagtg aagcttcgat gatgggctta ctgaccaacc
     1081 tggcagacag ggagctggtt cacatgatca actgggcgaa gagggtgcca ggctttgtgg
     1141 atttgaccct ccatgatcag gtccaccttc tagaatgtgc ctggctagag atcctgatga
     1201 ttggtctcgt ctggcgctcc atggagcacc cagtgaagct actgtttgct cctaacttgc
     1261 tcttggacag gaaccaggga aaatgtgtag agggcatggt ggagatcttc gacatgctgc
     1321 tggctacatc atctcggttc cgcatgatga atctgcaggg agaggagttt gtgtgcctca
     1381 aatctattat tttgcttaat tctggagtgt acacatttct gtccagcacc ctgaagtctc
     1441 tggaagagaa ggaccatatc caccgagtcc tggacaagat cacagacact ttgatccacc
     1501 tgatggccaa ggcaggcctg accctgcagc agcagcacca gcggctggcc cagctcctcc
     1561 tcatcctctc ccacatcagg cacatgagta acaaaggcat ggagcatctg tacagcatga
     1621 agtgcaagaa cgtggtgccc ctctatgacc tgctgctgga ggcggcggac gcccaccgcc
     1681 tacatgcgcc cactagccgt ggaggggcat ccgtggagga gacggaccaa agccacttgg
     1741 ccactgcggg ctctacttca tcgcattcct tgcaaaagta ttacatcacg ggggaggcag
     1801 agggtttccc tgccacagct gggggtggtg gatcaaccgg tgaaatgatg ctgaaaaaaa
     1861 tcctgaaaat cgaggaactg gatgaacgcg gtaccggcag cggaggtcat accttgtatg
     1921 cgccgggggg ttatgacatc atgggttacc tcatacagat catgaatagg ccgaacccac
     1981 aagtggaact cggacccgtc gatacctcct gcgctctcat tctgtgtgac cttaagcaga
     2041 aggatacccc tatcgtgtac gcctccgagg catttctgta catgacaggg tactcgaacg
     2101 ccgaagtgct gggacggaac tgccgcttcc tgcaaagccc ggatggaatg gtgaagccta
     2161 agtcaacccg caaatacgtg gactccaaca ctatcaacac catgcgcaag gccattgacc
     2221 gcaatgctga ggtgcaagtg gaagtggtga acttcaagaa gaatggacag cgcttcgtca
     2281 acttcctgac tatgatcccc gtgcgcgacg agaccggcga atatcggtac agcatggggt
     2341 ttcagtgtga gacagagggc ggctcaggag gcagcggcgg ttctggaggt tccggtggcg
     2401 gctcaggagg tagcggaggg tctcacactc tttacgcccc tggaggatac gacattatgg
     2461 gatatttgat tcagattatg aaccgcccaa accctcaggt cgaactgggg cctgtggaca
     2521 cgtcatgtgc cctgatcctg tgcgatctga agcaaaagga cactccgatt gtctacgcct
     2581 cggaagcctt cttgtatatg accggataca gcaatgcaga ggtgctcggt aggaactgca
     2641 gattcctgca gtcccccgac gggatggtga aaccaaagtc cactcgcaaa tatgtggact
     2701 cgaacacgat caatacaatg cggaaggcca tcgaccggaa cgccgaggtc caggtggagg
     2761 tggtcaactt taagaagaac ggccagcggt tcgtgaactt tctcaccatg attccggtcc
     2821 gggatgaaac cggagagtac agatactcca tgggattcca gtgcgaaacc gaagggtcag
     2881 gatccgaact gattgatatt gaagttagcg gtaaccacct gttctatgcc aatgatattc
     2941 tgacccataa tagctcgagc tctagattag ataaaagtaa agtgattaac agcgcattag
     3001 agctgcttaa tgaggtcgga atcgaaggtt taacaacccg taaactcgcc cagaagctag
     3061 gtgtagagca gcctacattg tattggcatg taaaaaataa gcgggctttg ctcgacgcct
     3121 tagccattga gatgttagat aggcaccata ctcacttttg ccctttagaa ggggaaagct
     3181 ggcaagattt tttacgtaat aacgctaaaa gttttagatg tgctttacta agtcatcgcg
     3241 atggagcaaa agtacattta ggtacacggc ctacagaaaa acagtatgaa actctcgaaa
     3301 atcaattagc ctttttatgc caacaaggtt tttcactaga gaatgcatta tatgcactca
     3361 gcgctgtggg gcattttact ttaggttgcg tattggaaga tcaagagcat caagtcgcta
     3421 aagaagaaag ggaaacacct actactgata gtatgccgcc attattacga caagctatcg
     3481 aattatttga tcaccaaggt gcagagccag ccttcttatt cggccttgaa ttgatcatat
     3541 gcggattaga aaaacaactt aaatgtgaaa gtgggtccgc gtacagccgc gcgcgtacga
     3601 aaaacaatta cgggtctacc atcgagggcc tgctcgatct cccggacgac gacgcccccg
     3661 aagaggcggg gctggcggct ccgcgcctgt cctttctccc cgcgggacac acgcgcagac
     3721 tgtcgacggc ccccccgacc gatgtcagcc tgggggacga gctccactta gacggcgagg
     3781 acgtggcgat ggcgcatgcc gacgcgctag acgatttcga tctggacatg ttgggggacg
     3841 gggattcccc gggtccggga tttacccccc acgactccgc cccctacggc gctctggata
     3901 tggccgactt cgagtttgag cagatgttta ccgatgccct tggaattgac gagtacggtg
     3961 ggtaaagcgg ccgcgactct agatcataat cagccatacc acatttgtag aggttttact
     4021 tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt
     4081 tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     4141 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     4201 tgtatcttaa ggcgtaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt
     4261 gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa
     4321 aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa
     4381 agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac
     4441 gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga
     4501 accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa
     4561 aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc
     4621 tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg tcaggtggca
     4681 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     4741 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     4801 gtcctgaggc ggaaagaacc agctgtggaa tgtgtgtcag ttagggtgtg gaaagtcccc
     4861 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccaggtg
     4921 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     4981 agcaaccata gtcccgcccc taactccgcc catcccgccc ctaactccgc ccagttccgc
     5041 ccattctccg ccccatggct gactaatttt ttttatttat gcagaggccg aggccgcctc
     5101 ggcctctgag ctattccaga agtagtgagg aggctttttt ggaggcctag gcttttgcaa
     5161 agatcgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg
     5221 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
     5281 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
     5341 tcaagaccga cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt
     5401 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
     5461 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
     5521 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
     5581 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
     5641 aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg
     5701 aactgttcgc caggctcaag gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg
     5761 gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact
     5821 gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg
     5881 ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc
     5941 ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga gcgggactct
     6001 ggggttcgaa atgaccgacc aagcgacgcc caacctgcca tcacgagatt tcgattccac
     6061 cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat
     6121 cctccagcgc ggggatctca tgctggagtt cttcgcccac cctaggggga ggctaactga
     6181 aacacggaag gagacaatac cggaaggaac ccgcgctatg acggcaataa aaagacagaa
     6241 taaaacgcac ggtgttgggt cgtttgttca taaacgcggg gttcggtccc agggctggca
     6301 ctctgtcgat accccaccga gaccccattg gggccaatac gcccgcgttt cttccttttc
     6361 cccaccccac cccccaagtt cgggtgaagg cccagggctc gcagccaacg tcggggcggc
     6421 aggccctgcc atagcctcag gttactcata tatactttag attgatttaa aacttcattt
     6481 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     6541 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     6601 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     6661 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     6721 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     6781 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     6841 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     6901 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     6961 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     7021 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     7081 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     7141 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     7201 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     7261 atcccctgat tctgtggata accgtattac cgccatgcat tagttattaa tagtaatcaa
     7321 ttacggggtc attagttcat agcccatata tggagttccg
//
