LOCUS       sequence-319793-        5238 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 5238)
  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 5238)
  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: mRuby-PH-PLCdelta
FEATURES             Location/Qualifiers
     source          1..5238
                     /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      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             613..1323
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTH
                     MALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAV
                     AKFAGLGGGMDELYK"
     misc_feature    1324..1380
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    2026..2147
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2063..2082)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2117..2136
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2154..2609)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2291..2312)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2503..2522
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2636..2740
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2742..3099
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2742..2933
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2765..2785)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2950..3085
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3134..3928
                     /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             3134..3928
                     /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(3188..3207)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3798..3817
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4116..4135)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4160..4207
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4536..5124
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5025..5044
                     /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 gctagcgcta
      601 ccggtcgcca ccatggtgtc taagggcgaa gagctgatca aggaaaatat gcgtatgaag
      661 gtggtcatgg aaggttcggt caacggccac caattcaaat gcacaggtga aggagaaggc
      721 aatccgtaca tgggaactca aaccatgagg atcaaagtca tcgagggagg acccctgcca
      781 tttgcctttg acattcttgc cacgtcgttc atgtatggca gccgtacttt tatcaagtac
      841 ccgaaaggca ttcctgattt ctttaaacag tcctttcctg agggttttac ttgggaaaga
      901 gttacgagat acgaagatgg tggagtcgtc accgtcatgc aggacaccag ccttgaggat
      961 ggctgtctcg tttaccacgt ccaagtcaga ggggtaaact ttccctccaa tggtcccgtg
     1021 atgcagaaga agaccaaggg ttgggagcct aatacagaga tgatgtatcc agcagatggt
     1081 ggtctgaggg gatacactca tatggcactg aaagttgatg gtggtggcca tctgtcttgc
     1141 tctttcgtaa caacttacag gtcaaaaaag accgtcggga acatcaagat gcccggtatc
     1201 catgccgttg atcaccgcct ggaaaggtta gaggaaagtg acaatgaaat gttcgtagta
     1261 caacgcgaac acgcagttgc caagttcgcc gggcttggtg gtgggatgga cgagctgtac
     1321 aagagatctc gagctcaagc ttcgaattct gcagtcgacg gtaccgcggg cccgggatcc
     1381 atggactcgg gccgggactt cctgaccctg cacggcctac aggatgatga ggatctacag
     1441 gcgctgctga agggcagcca gctcctgaag gtgaagtcca gctcatggag gagagagcgc
     1501 ttctacaagt tgcaggagga ctgcaagacc atctggcagg agtcccgcaa ggtcatgcgg
     1561 accccggagt cccagctgtt ctccatcgag gacattcagg aggtgcgaat ggggcaccgc
     1621 acggagggtc tggagaagtt cgcccgtgat gtgcccgagg accgctgctt ctccattgtc
     1681 ttcaaggacc agcgcaatac actagacctc atcgccccat cgccagctga tgcccagcac
     1741 tgggtgctgg ggctgcacaa gatcatccac cactcaggct ccatggacca gcgtcagaag
     1801 ctacagcact ggattcactc ctgcttgcga aaagctgaca aaaacaagga caacaagatg
     1861 agcttcaagg agctgcagaa cttcctgaag gagctcaaca tccagtaatc tagataactg
     1921 atcataatca gccataccac atttgtagag gttttacttg ctttaaaaaa cctcccacac
     1981 ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt gtttattgca
     2041 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt
     2101 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttaacg cgtaaattgt
     2161 aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatcagct cattttttaa
     2221 ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt
     2281 gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa
     2341 agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag
     2401 ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt
     2461 tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg
     2521 agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc
     2581 cgcgcttaat gcgccgctac agggcgcgtc aggtggcact tttcggggaa atgtgcgcgg
     2641 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     2701 accctgataa atgcttcaat aatattgaaa aaggaagagt cctgaggcgg aaagaaccag
     2761 ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt
     2821 atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca
     2881 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta
     2941 actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga
     3001 ctaatttttt ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag
     3061 tagtgaggag gcttttttgg aggcctaggc ttttgcaaag atcgatcaag agacaggatg
     3121 aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt
     3181 ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt
     3241 gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc
     3301 cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc
     3361 ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga
     3421 agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat
     3481 ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca
     3541 agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga
     3601 tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc
     3661 gagcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat
     3721 catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga
     3781 ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg
     3841 ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt
     3901 ctatcgcctt cttgacgagt tcttctgagc gggactctgg ggttcgaaat gaccgaccaa
     3961 gcgacgccca acctgccatc acgagatttc gattccaccg ccgccttcta tgaaaggttg
     4021 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     4081 ctggagttct tcgcccaccc tagggggagg ctaactgaaa cacggaagga gacaataccg
     4141 gaaggaaccc gcgctatgac ggcaataaaa agacagaata aaacgcacgg tgttgggtcg
     4201 tttgttcata aacgcggggt tcggtcccag ggctggcact ctgtcgatac cccaccgaga
     4261 ccccattggg gccaatacgc ccgcgtttct tccttttccc caccccaccc cccaagttcg
     4321 ggtgaaggcc cagggctcgc agccaacgtc ggggcggcag gccctgccat agcctcaggt
     4381 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     4441 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     4501 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     4561 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     4621 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     4681 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     4741 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     4801 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     4861 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     4921 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     4981 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     5041 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     5101 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     5161 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     5221 cgtattaccg ccatgcat
//
