LOCUS       sequence-319788-        4904 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 4904)
  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 4904)
  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-FRB2C
FEATURES             Location/Qualifiers
     source          1..4904
                     /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    1692..1813
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1729..1748)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1783..1802
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(1820..2275)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1957..1978)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2169..2188
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2302..2406
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2408..2765
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2408..2599
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2431..2451)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2616..2751
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             2800..3594
                     /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             2800..3594
                     /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(2854..2873)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3464..3483
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(3782..3801)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    3826..3873
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4202..4790
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4691..4710
                     /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 ggtaccaatg
     1501 tcaaggacct cctccaagcc tgggacctct attatcatgt gttccgacga atctcaaagt
     1561 agggatccac cggatctaga taactgatca taatcagcca taccacattt gtagaggttt
     1621 tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa
     1681 ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca
     1741 caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca
     1801 tcaatgtatc ttaaggcgta aattgtaagc gttaatattt tgttaaaatt cgcgttaaat
     1861 ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat cccttataaa
     1921 tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa gagtccacta
     1981 ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca
     2041 ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat
     2101 cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg
     2161 agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc
     2221 acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcaggt
     2281 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     2341 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     2401 aagagtcctg aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt
     2461 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     2521 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
     2581 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
     2641 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
     2701 cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
     2761 gcaaagatcg atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg
     2821 cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag
     2881 acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt
     2941 tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aagacgaggc agcgcggcta
     3001 tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg
     3061 ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt
     3121 gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat
     3181 ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg
     3241 atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca
     3301 gccgaactgt tcgccaggct caaggcgagc atgcccgacg gcgaggatct cgtcgtgacc
     3361 catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc
     3421 gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat
     3481 attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc
     3541 gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga
     3601 ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt
     3661 ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga
     3721 tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccctagg gggaggctaa
     3781 ctgaaacacg gaaggagaca ataccggaag gaacccgcgc tatgacggca ataaaaagac
     3841 agaataaaac gcacggtgtt gggtcgtttg ttcataaacg cggggttcgg tcccagggct
     3901 ggcactctgt cgatacccca ccgagacccc attggggcca atacgcccgc gtttcttcct
     3961 tttccccacc ccacccccca agttcgggtg aaggcccagg gctcgcagcc aacgtcgggg
     4021 cggcaggccc tgccatagcc tcaggttact catatatact ttagattgat ttaaaacttc
     4081 atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc
     4141 cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt
     4201 cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac
     4261 cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct
     4321 tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta ggccaccact
     4381 tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg
     4441 ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata
     4501 aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga
     4561 cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag
     4621 ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg
     4681 agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac
     4741 ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca
     4801 acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg
     4861 cgttatcccc tgattctgtg gataaccgta ttaccgccat gcat
//
