LOCUS       sequence-199266-        6269 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  'Localizer' construct that marks lacO arrays and is targeted by a 
            PHR-tagged effector upon illumination with blue light; can be 
            visualized without triggering PHR recruitment.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6269)
  AUTHORS   Rademacher A, Erdel F, Trojanowski J, Schumacher S, Rippe K
  TITLE     Real-time observation of light-controlled transcription in living 
            cells.
  JOURNAL   J Cell Sci. 2017 Dec 15;130(24):4213-4224. doi: 10.1242/jcs.205534. 
            Epub 2017 Nov 9.
  PUBMED    29122982
REFERENCE   2  (bases 1 to 6269)
  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: 
            "10.1242/jcs.205534"; journalName: "J Cell Sci"; date: "2017-12-15- 
            15"; volume: "130"; issue: "24"; pages: "4213-4224"
COMMENT     Sequence Label: pCIBN-LacI-tagRFP-T
FEATURES             Location/Qualifiers
     source          1..6269
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        59..362
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        363..566
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     517..537
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             601..1110
                     /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             1126..2205
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(1145..1164)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     CDS             2212..2925
                     /codon_start=1
                     /product="monomeric derivative of red fluorescent protein 
                     from Entacmaea quadricolor (Merzlyak et al., 2007)"
                     /label=TagRFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTM
                     RIKVVEGGPLPFAFDILATSFMYGSRTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGG
                     VLTATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEANTEMLYPADGGLEGRTD
                     MALKLVGGGHLICNFKTTYRSKKPAKNLKMPGVYYVDHRLERIKEADKETYVEQHEVAV
                     ARYCDLPSKLGHKLN"
     polyA_signal    3055..3176
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3092..3111)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3146..3165
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3183..3638)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3320..3341)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3532..3551
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3665..3769
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3771..4128
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3771..3962
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3794..3814)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3979..4114
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4163..4957
                     /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             4163..4957
                     /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(4217..4236)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4827..4846
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5145..5164)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5189..5236
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5565..6153
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6054..6073
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gttattaata gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg
       61 ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga
      121 cgtcaataat gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat
      181 gggtggagta tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa
      241 gtacgccccc tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca
      301 tgaccttatg ggactttcct acttggcagt acatctacgt attagtcatc gctattacca
      361 tggtgatgcg gttttggcag tacatcaatg ggcgtggata gcggtttgac tcacggggat
      421 ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg
      481 actttccaaa atgtcgtaac aactccgccc cattgacgca aatgggcggt aggcgtgtac
      541 ggtgggaggt ctatataagc agagctggtt tagtgaaccg tcagatccgc tagcgccacc
      601 atgaatggag ctataggagg tgaccttttg ctcaattttc ctgacatgtc ggtcctagag
      661 cgccaaaggg ctcacctcaa gtacctcaat cccacctttg attctcctct cgccggcttc
      721 tttgccgatt cttcaatgat taccggcggc gagatggaca gctatctttc gactgccggt
      781 ttgaatcttc cgatgatgta cggtgagacg acggtggaag gtgattcaag actctcaatt
      841 tcgccggaaa cgacgcttgg gactggaaat ttcaaggcag cgaagtttga tacagagact
      901 aaggattgta atgaggcggc gaagaagatg acgatgaaca gagatgacct agtagaagaa
      961 ggagaagaag agaagtcgaa aataacagag caaaacaatg ggagcacaaa aagcatcaag
     1021 aagatgaaac acaaagccaa gaaagaagag aacaatttct ctaatgattc atctaaagtg
     1081 acgaaggaat tggagaaaac ggattatatt catgtaccgg taatggtgaa accagtaacg
     1141 ttatacgatg tcgcagagta tgccggtgtc tcttatcaga ccgtttcccg cgtggtgaac
     1201 caggccagcc acgtttctgc gaaaacgcgg gaaaaagtgg aagcggcgat ggcggagctg
     1261 aattacattc ccaaccgcgt ggcacaacaa ctggcgggca aacagtcgtt gctgattggc
     1321 gttgccacct ccagtctggc cctgcacgcg ccgtcgcaaa ttgtcgcggc gattaaatct
     1381 cgcgccgatc aactgggtgc cagcgtggtg gtgtcgatgg tagaacgaag cggcgtcgaa
     1441 gcctgtaaag cggcggtgca caatcttctc gcgcaacgcg tcagtgggct gatcattaac
     1501 tatccgctgg atgaccagga tgccattgct gtggaagctg cctgcactaa tgttccggcg
     1561 ttatttcttg atgtctctga ccagacaccc atcaacagta ttattttctc ccatgaagac
     1621 ggtacgcgac tgggcgtgga gcatctggtc gcattgggtc accagcaaat cgcgctgtta
     1681 gcgggcccat taagttctgt ctcggcgcgt ctgcgtctgg ctggctggca taaatatctc
     1741 actcgcaatc aaattcagcc gatagcggaa cgggaaggcg actggagtgc catgtccggt
     1801 tttcaacaaa ccatgcaaat gctgaatgag ggcatcgttc ccactgcgat gctggttgcc
     1861 aacgatcaga tggcgctggg cgcaatgcgc gccattaccg agtccgggct gcgcgttggt
     1921 gcggatatct cggtagtggg atacgacgat accgaagaca gctcatgtta tatcccgccg
     1981 ttaaccacca tcaaacagga ttttcgcctg ctggggcaaa ccagcgtgga ccgcttgctg
     2041 caactctctc agggccaggc ggtgaagggc aatcagctgt tgcccgtctc actggtgaaa
     2101 agaaaaacca ccctggcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca
     2161 ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg ggcagaagct tatggtgtct
     2221 aagggcgaag agctgattaa ggagaacatg cacatgaagc tgtacatgga gggcaccgtg
     2281 aacaaccacc acttcaagtg cacatccgag ggcgaaggca agccctacga gggcacccag
     2341 accatgagaa tcaaggtggt cgagggcggc cctctcccct tcgccttcga catcctggct
     2401 accagcttca tgtacggcag cagaaccttc atcaaccaca cccagggcat ccccgacttc
     2461 tttaagcagt ccttccctga gggcttcaca tgggagagag tcaccacata cgaagacggg
     2521 ggcgtgctga ccgctaccca ggacaccagc ctccaggacg gctgcctcat ctacaacgtc
     2581 aagatcagag gggtgaactt cccatccaac ggccctgtga tgcagaagaa aacactcggc
     2641 tgggaggcca acaccgagat gctgtacccc gctgacggcg gcctggaagg cagaaccgac
     2701 atggccctga agctcgtggg cgggggccac ctgatctgca acttcaagac cacatacaga
     2761 tccaagaaac ccgctaagaa cctcaagatg cccggcgtct actatgtgga ccacagactg
     2821 gaaagaatca aggaggccga caaagagacc tacgtcgagc agcacgaggt ggctgtggcc
     2881 agatactgcg acctccctag caaactgggg cacaaactta attaaggatc caccggatct
     2941 agataactga tcataatcag ccataccaca tttgtagagg ttttacttgc tttaaaaaac
     3001 ctcccacacc tccccctgaa cctgaaacat aaaatgaatg caattgttgt tgttaacttg
     3061 tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa
     3121 gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttaacgc
     3181 gtaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
     3241 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
     3301 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
     3361 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
     3421 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
     3481 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
     3541 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
     3601 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa
     3661 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     3721 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagtc ctgaggcgga
     3781 aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     3841 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     3901 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     3961 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     4021 catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta
     4081 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaga tcgatcaaga
     4141 gacaggatga ggatcgtttc gcatgattga acaagatgga ttgcacgcag gttctccggc
     4201 cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga
     4261 tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct
     4321 gtccggtgcc ctgaatgaac tgcaagacga ggcagcgcgg ctatcgtggc tggccacgac
     4381 gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct
     4441 attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt
     4501 atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt
     4561 cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt
     4621 cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag
     4681 gctcaaggcg agcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt
     4741 gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg
     4801 tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg
     4861 cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg
     4921 catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg
     4981 accgaccaag cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat
     5041 gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg
     5101 gatctcatgc tggagttctt cgcccaccct agggggaggc taactgaaac acggaaggag
     5161 acaataccgg aaggaacccg cgctatgacg gcaataaaaa gacagaataa aacgcacggt
     5221 gttgggtcgt ttgttcataa acgcggggtt cggtcccagg gctggcactc tgtcgatacc
     5281 ccaccgagac cccattgggg ccaatacgcc cgcgtttctt ccttttcccc accccacccc
     5341 ccaagttcgg gtgaaggccc agggctcgca gccaacgtcg gggcggcagg ccctgccata
     5401 gcctcaggtt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     5461 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     5521 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     5581 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     5641 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     5701 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     5761 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     5821 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     5881 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     5941 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     6001 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     6061 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     6121 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     6181 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     6241 gtggataacc gtattaccgc catgcatta
//
