LOCUS       sequence-351702-        5153 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of a protein of interest fused to the 
            C-terminus of miRFP670-Zdk1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5153)
  AUTHORS   Yoshikawa M, Yoshii T, Ikuta M, Tsukiji S
  TITLE     Synthetic Protein Condensates That Inducibly Recruit and Release 
            Protein Activity in Living Cells.
  JOURNAL   J Am Chem Soc. 2021 May 5;143(17):6434-6446. doi: 
            10.1021/jacs.0c12375. Epub 2021 Apr 23.
  PUBMED    33890764
REFERENCE   2  (bases 1 to 5153)
  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: 
            "10.1021/jacs.0c12375"; journalName: "J Am Chem Soc"; date: 
            "2021-05-5- 5"; volume: "143"; issue: "17"; pages: "6434-6446"
COMMENT     Sequence Label: pCMV-miRFP670-Zdk1-MCS
FEATURES             Location/Qualifiers
     source          1..5153
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        56..359
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        360..563
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     514..534
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      602..611
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             608..1552
                     /codon_start=1
                     /product="bright monomeric near-infrared fluorescent 
                     protein engineered from a bacterial phytochrome, with an 
                     emission maximum of 670 nm (Shcherbakova et al., 2016)"
                     /label=miRFP670
                     /label=ORF frame 1
                     /translation="MVAGHASGSPAFGTASHSNCEHEEIHLAGSIQPHGALLVVSEHDH
                     RVIQASANAAEFLNLGSVLGVPLAEIDGDLLIKILPHLDPTAEGMPVAVRCRIGNPSTE
                     YCGLMHRPPEGGLIIELERAGPSIDLSGTLAPALERIRTAGSLRALCDDTVLLFQQCTG
                     YDRVMVYRFDEQGHGLVFSECHVPGLESYFGNRYPSSTVPQMARQLYVRQRVRVLVDVT
                     YQPVPLEPRLSPLTGRDLDMSGCFLRSMSPCHLQFLKDMGVRATLAVSLVVGGKLWGLV
                     VCHHYLPRFIRFELRAICKRLAERIATRITALES"
     CDS             1583..1759
                     /codon_start=1
                     /product="""Zdark"" protein, derived from the Z subunit of 
                     protein A, that binds to the dark state of LOV2 and is 
                     released upon irradiation (Wang et al., 2016)"
                     /label=Zdk1
                     /translation="MVDNKFNKEKTRAGAEIHSLPNLNVEQKFAFIVSLFDDPSQSANL
                     LAEAKKLNDAQAPK"
     polyA_signal    1936..2057
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1973..1992)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2027..2046
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2064..2519)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2201..2222)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2413..2432
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        2546..2650
                     /gene="bla"
                     /label=AmpR promoter
     promoter        2652..3009
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        2652..2843
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(2675..2695)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      2860..2995
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3044..3838
                     /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             3044..3838
                     /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(3098..3117)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3708..3727
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4026..4045)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4070..4117
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      4446..5034
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4935..4954
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 attaatagta atcaattacg gggtcattag ttcatagccc atatatggag ttccgcgtta
       61 cataacttac ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt
      121 caataatgac gtatgttccc atagtaacgc caatagggac tttccattga cgtcaatggg
      181 tggagtattt acggtaaact gcccacttgg cagtacatca agtgtatcat atgccaagta
      241 cgccccctat tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc cagtacatga
      301 ccttatggga ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgg
      361 tgatgcggtt ttggcagtac atcaatgggc gtggatagcg gtttgactca cggggatttc
      421 caagtctcca ccccattgac gtcaatggga gtttgttttg gcaccaaaat caacgggact
      481 ttccaaaatg tcgtaacaac tccgccccat tgacgcaaat gggcggtagg cgtgtacggt
      541 gggaggtcta tataagcaga gctggtttag tgaaccgtca gatccgctag cgctaccggt
      601 cgccaccatg gtagcaggtc atgcctctgg cagccccgca ttcgggaccg cctctcattc
      661 gaattgcgaa catgaagaga tccacctcgc cggctcgatc cagccgcatg gcgcgcttct
      721 ggtcgtcagc gaacatgatc atcgcgtcat ccaggccagc gccaacgccg cggaatttct
      781 gaatctcgga agcgtactcg gcgttccgct cgccgagatc gacggcgatc tgttgatcaa
      841 gatcctgccg catctcgatc ccaccgccga aggcatgccg gtcgcggtgc gctgccggat
      901 cggcaatccc tctacggagt actgcggtct gatgcatcgg cctccggaag gcgggctgat
      961 catcgaactc gaacgtgccg gcccgtcgat cgatctgtca ggcacgctgg cgccggcgct
     1021 ggagcggatc cgcacggcgg gttcactgcg cgcgctgtgc gatgacaccg tgctgctgtt
     1081 tcagcagtgc accggctacg accgggtgat ggtgtatcgt ttcgatgagc aaggccacgg
     1141 cctggtattc tccgagtgcc atgtgcctgg gctcgaatcc tatttcggca accgctatcc
     1201 gtcgtcgact gtcccgcaga tggcgcggca gctgtacgtg cggcagcgcg tccgcgtgct
     1261 ggtcgacgtc acctatcagc cggtgccgct ggagccgcgg ctgtcgccgc tgaccgggcg
     1321 cgatctcgac atgtcgggct gcttcctgcg ctcgatgtcg ccgtgccatc tgcagttcct
     1381 gaaggacatg ggcgtgcgcg ccaccctggc ggtgtcgctg gtggtcggcg gcaagctgtg
     1441 gggcctggtt gtctgtcacc attatctgcc gcgcttcatc cgtttcgagc tgcgggcgat
     1501 ctgcaaacgg ctcgccgaaa ggatcgcgac gcggatcacc gcgcttgaga gcctgtacaa
     1561 gtccggactc agatctcgag ctatggtgga taacaaattc aataaagaaa agacgcgtgc
     1621 cggtgcggaa atccattctc tgccaaacct gaatgttgag cagaagtttg ccttcatcgt
     1681 gagcctgttt gatgatccat ctcagagcgc caatctgctg gccgaagcca aaaaactgaa
     1741 cgatgcccag gccccaaaaa ctagttaaaa gcttcgaatt ctgcagtcga cggtaccgcg
     1801 ggcccgggat ccaccggatc tagataactg atcataatca gccataccac atttgtagag
     1861 gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat
     1921 gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     1981 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     2041 ctcatcaatg tatcttaagg cgtaaattgt aagcgttaat attttgttaa aattcgcgtt
     2101 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta
     2161 taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc
     2221 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg
     2281 cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact
     2341 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt
     2401 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc
     2461 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc
     2521 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     2581 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     2641 aaggaagagt cctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga
     2701 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     2761 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     2821 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     2881 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     2941 gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     3001 ttttgcaaag atcgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg
     3061 attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca
     3121 acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt
     3181 tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg
     3241 gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga
     3301 agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca
     3361 ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct
     3421 tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac
     3481 tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc
     3541 gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt
     3601 gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt
     3661 catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg
     3721 tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat
     3781 cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc
     3841 gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc acgagatttc
     3901 gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc
     3961 tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc tagggggagg
     4021 ctaactgaaa cacggaagga gacaataccg gaaggaaccc gcgctatgac ggcaataaaa
     4081 agacagaata aaacgcacgg tgttgggtcg tttgttcata aacgcggggt tcggtcccag
     4141 ggctggcact ctgtcgatac cccaccgaga ccccattggg gccaatacgc ccgcgtttct
     4201 tccttttccc caccccaccc cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc
     4261 ggggcggcag gccctgccat agcctcaggt tactcatata tactttagat tgatttaaaa
     4321 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     4381 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     4441 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     4501 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     4561 ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac
     4621 cacttcaaga actctgtatc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     4681 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     4741 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     4801 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     4861 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     4921 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     4981 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     5041 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     5101 cctgcgttat cccctgattc tgtggataac cgtattaccg ccatgcatta gtt
//
