LOCUS       sequence-319786-        4967 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  for use in CIT (chemically induced trimerization) system.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4967)
  AUTHORS   Wu HD, Kikuchi M, Dagliyan O, Aragaki AK, Nakamura H, Dokholyan NV, 
            Umehara T, Inoue T
  TITLE     Rational design and implementation of a chemically inducible 
            heterotrimerization system.
  JOURNAL   Nat Methods. 2020 Sep;17(9):928-936. doi: 10.1038/s41592-020-0913-x.
            Epub 2020 Aug 3.
  PUBMED    32747768
REFERENCE   2  (bases 1 to 4967)
  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: 
            "10.1038/s41592-020-0913-x"; journalName: "Nat Methods"; date: 
            "2020-09"; volume: "17"; issue: "9"; pages: "928-936"
COMMENT     Sequence Label: Lyn-YFP-FRB1N
FEATURES             Location/Qualifiers
     source          1..4967
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      597..606
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      663..672
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(669..671,672..674,675..1385)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(714..735)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(975..994)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1322..1343
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1755..1876
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1792..1811)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1846..1865
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1883..2338)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2020..2041)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2232..2251
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2365..2469
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2471..2828
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2471..2662
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2494..2514)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2679..2814
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2863..3657
                     /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             2863..3657
                     /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(2917..2936)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3527..3546
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3845..3864)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3889..3936
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4265..4853
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4754..4773
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcca
      601 ccatgggatg tataaaatca aaagggaaag acagcgcgga tagtgctggt ggtttaccgg
      661 tcgccaccat ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
      721 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg
      781 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
      841 ggcccaccct cgtgaccacc ttcggctacg gcctgcagtg cttcgcccgc taccccgacc
      901 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
      961 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
     1021 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     1081 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc
     1141 agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc
     1201 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     1261 acaaccacta cctgagctac cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc
     1321 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt
     1381 acaagtccgg actcagatct cgagctcaag cttcgaattc tgcagtcgac agtgctggtg
     1441 gtagtgctgg tggtagtgct ggtggtagtg ctggtggtag tgctggtggt ggtaccatcc
     1501 tctggcatga gatgtggcat gaaggcctgg aagaggcatc tcgtttgtac tttggggaaa
     1561 ggaacgtgaa aggcatgttt gaggtgctgg agcccttgca tgctatgatg gaacggggcc
     1621 cctagggatc caccggatct agataactga tcataatcag ccataccaca tttgtagagg
     1681 ttttacttgc tttaaaaaac ctcccacacc tccccctgaa cctgaaacat aaaatgaatg
     1741 caattgttgt tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     1801 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     1861 tcatcaatgt atcttaaggc gtaaattgta agcgttaata ttttgttaaa attcgcgtta
     1921 aatttttgtt aaatcagctc attttttaac caataggccg aaatcggcaa aatcccttat
     1981 aaatcaaaag aatagaccga gatagggttg agtgttgttc cagtttggaa caagagtcca
     2041 ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc
     2101 ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg taaagcacta
     2161 aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc ggcgaacgtg
     2221 gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg
     2281 gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca
     2341 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     2401 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     2461 aggaagagtc ctgaggcgga aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa
     2521 agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa
     2581 ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     2641 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     2701 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     2761 ccgcctcggc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     2821 tttgcaaaga tcgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga
     2881 ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa
     2941 cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt
     3001 ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaagacga ggcagcgcgg
     3061 ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa
     3121 gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac
     3181 cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt
     3241 gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact
     3301 cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg
     3361 ccagccgaac tgttcgccag gctcaaggcg agcatgcccg acggcgagga tctcgtcgtg
     3421 acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc
     3481 atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt
     3541 gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc
     3601 gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg
     3661 ggactctggg gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg
     3721 attccaccgc cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct
     3781 ggatgatcct ccagcgcggg gatctcatgc tggagttctt cgcccaccct agggggaggc
     3841 taactgaaac acggaaggag acaataccgg aaggaacccg cgctatgacg gcaataaaaa
     3901 gacagaataa aacgcacggt gttgggtcgt ttgttcataa acgcggggtt cggtcccagg
     3961 gctggcactc tgtcgatacc ccaccgagac cccattgggg ccaatacgcc cgcgtttctt
     4021 ccttttcccc accccacccc ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg
     4081 gggcggcagg ccctgccata gcctcaggtt actcatatat actttagatt gatttaaaac
     4141 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     4201 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     4261 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     4321 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     4381 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     4441 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     4501 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     4561 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     4621 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     4681 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     4741 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     4801 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     4861 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
     4921 ctgcgttatc ccctgattct gtggataacc gtattaccgc catgcat
//
