LOCUS       sequence-474341-        5300 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the I3-01 nanocage subunit N-terminally tagged with 
            mStayGold(E138D) in mammalian cells. Assembles into nanocages tagged
            with 60 FPs..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5300)
  AUTHORS   Viola G, Ibrahim YW, Jacobs KA, Lemiere J, Kutys ML, Wittmann T
  TITLE     Quantitative comparison of fluorescent proteins using protein 
            nanocages in live cells.
  JOURNAL   J Cell Sci. 2025 May 15;138(10):jcs263858. doi: 10.1242/jcs.263858. 
            Epub 2025 May 21.
  PUBMED    40326462
REFERENCE   2  (bases 1 to 5300)
  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"; journalName: "J Cell 
            Sci."; date: "2025-05-15"; volume: "138(10):jcs263858. doi"; pages: 
            " 10.1242/jcs.263858. Epub 2025 May 21"
COMMENT     Sequence Label: pmStayGold(E138D)_I3
FEATURES             Location/Qualifiers
     source          1..5300
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        764..1067
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1068..1271
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1222..1242
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    1312..1368
                     /label=MCS
                     /note="multiple cloning site"
     regulatory      1377..1386
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1383..2033
                     /codon_start=1
                     /product="bright and highly photostable monomeric green 
                     fluorescent protein (Ivorra-Molla et al., 2023)"
                     /label=mStayGold (E138D)
                     /note="contains the monomerizing E138D mutation"
                     /translation="MASTPFKFQLKGTINGKSFTVEGEGEGNSHEGSHKGKYVCTSGKL
                     PMSWAALGTSFGYGMKYYTKYPSGLKNWFHEVMPEGFTYDRHIQYKGDGSIHAKHQHFM
                     KNGTYHNIVEFTGQDFKENSPVLTGDMNVSLPNDVQHIPRDDGVECPVTLLYPLLSDKS
                     KCVEAHQNTICKPLHNQPAPDVPYHWIRKQYTQSKDDTEERDHICQSETLEAHL"
     polyA_signal    2812..2933
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2849..2868)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2903..2922
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2940..3395)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3077..3098)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3289..3308
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3422..3526
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        3528..3719
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3551..3571)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3716..3850
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3899..4693
                     /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             3899..4693
                     /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(3953..3972)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4563..4582
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4881..4900)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4925..4972
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccatg cattagttat taatagtaat
      721 caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg
      781 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt
      841 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac
      901 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg
      961 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact
     1021 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt
     1081 ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc
     1141 ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc
     1201 gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata
     1261 taagcagagc tggtttagtg aaccgtcaga tccgctagcg ctaccggact cagatctcga
     1321 gctcaagctt cgaattctgc agtcgacggt accgcgggcc cgggatccaa ccggtcgcca
     1381 ccatggccag cacgccgttt aagttccagt taaaggggac aattaacggg aaaagcttta
     1441 cagtcgaggg tgaaggggaa gggaactcgc atgaaggtag tcataaaggt aagtatgtgt
     1501 gtacgagcgg aaagttgccg atgtcttggg cggctcttgg tacttcgttt gggtacggaa
     1561 tgaaatacta tacaaaatac ccgtcgggcc tgaaaaattg gttccatgag gtgatgcccg
     1621 aggggttcac gtatgatcgc cacatccagt acaaaggtga tggctcaatc cacgcaaagc
     1681 accaacactt catgaagaat gggacttatc ataacattgt cgagtttact ggccaggact
     1741 tcaaggaaaa cagccccgta ttgacaggtg acatgaatgt gagcttgccg aacgacgtgc
     1801 agcatattcc acgtgacgac ggagttgaat gtcccgtcac attgttatac cccttgctta
     1861 gcgataaatc aaagtgtgtc gaagcacatc aaaacactat ctgcaaacct cttcataacc
     1921 aacctgcgcc agatgtaccg taccattgga ttcgcaaaca atatacacag tctaaggatg
     1981 atacggagga acgcgatcat atctgtcagt ccgagacgtt agaagctcat cttggtgggt
     2041 caggcggatc cggtggtagc ggtggtagcg gtggtagcgg tggtagcgag gaactcttta
     2101 agaagcataa aatcgtagct gtgctcagag caaattcagt agaggaggcg aagaaaaagg
     2161 ccctggccgt tttcctgggc ggagtccacc tcatcgaaat tacttttact gtccccgacg
     2221 cagatacagt cataaaagag ttgagttttc tcaaagagat gggggcgatc atcggtgccg
     2281 ggacggttac gtcagtcgag cagtgccgga aagcagtgga gtcaggagcg gaatttatcg
     2341 tatctccaca tctggatgag gaaatatctc aattctgtaa agaaaaaggt gtcttttaca
     2401 tgcccggcgt tatgacgcca acggagctgg tgaaggccat gaagctggga cacacgatac
     2461 tggctctctt tccaggcgaa gtggtcggtc ctcaattcgt caaggcaatg aaagggcctt
     2521 tccccaacgt taaattcgtc ccgactgggg gagtaaactt ggacaatgtt tgcgaatggt
     2581 ttaaggcagg ggtactcgct gttggagttg gttccgcatt ggtaaaaggt acgcctgtcg
     2641 aagtggcaga aaaggcaaag gctttcgttg agaagatccg cggatgtact gaataggcga
     2701 ctctagatca taatcagcca taccacattt gtagaggttt tacttgcttt aaaaaacctc
     2761 ccacacctcc ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt taacttgttt
     2821 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
     2881 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttaaggcgta
     2941 aattgtaagc gttaatattt tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt
     3001 ttttaaccaa taggccgaaa tcggcaaaat cccttataaa tcaaaagaat agaccgagat
     3061 agggttgagt gttgttccag tttggaacaa gagtccacta ttaaagaacg tggactccaa
     3121 cgtcaaaggg cgaaaaaccg tctatcaggg cgatggccca ctacgtgaac catcacccta
     3181 atcaagtttt ttggggtcga ggtgccgtaa agcactaaat cggaacccta aagggagccc
     3241 ccgatttaga gcttgacggg gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc
     3301 gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc acgctgcgcg taaccaccac
     3361 acccgccgcg cttaatgcgc cgctacaggg cgcgtcaggt ggcacttttc ggggaaatgt
     3421 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
     3481 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtcctg aggcggaaag
     3541 aaccagctgt ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc
     3601 agaagtatgc aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc
     3661 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg
     3721 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     3781 tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg aggaggcttt
     3841 tttggaggcc taggcttttg caaagatcga tcaagagaca ggatgaggat cgtttcgcat
     3901 gattgaacaa gatggattgc acgcaggttc tccggccgct tgggtggaga ggctattcgg
     3961 ctatgactgg gcacaacaga caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc
     4021 gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc ggtgccctga atgaactgca
     4081 agacgaggca gcgcggctat cgtggctggc cacgacgggc gttccttgcg cagctgtgct
     4141 cgacgttgtc actgaagcgg gaagggactg gctgctattg ggcgaagtgc cggggcagga
     4201 tctcctgtca tctcaccttg ctcctgccga gaaagtatcc atcatggctg atgcaatgcg
     4261 gcggctgcat acgcttgatc cggctacctg cccattcgac caccaagcga aacatcgcat
     4321 cgagcgagca cgtactcgga tggaagccgg tcttgtcgat caggatgatc tggacgaaga
     4381 gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc aaggcgagca tgcccgacgg
     4441 cgaggatctc gtcgtgaccc atggcgatgc ctgcttgccg aatatcatgg tggaaaatgg
     4501 ccgcttttct ggattcatcg actgtggccg gctgggtgtg gcggaccgct atcaggacat
     4561 agcgttggct acccgtgata ttgctgaaga gcttggcggc gaatgggctg accgcttcct
     4621 cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc gccttctatc gccttcttga
     4681 cgagttcttc tgagcgggac tctggggttc gaaatgaccg accaagcgac gcccaacctg
     4741 ccatcacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt
     4801 ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc
     4861 caccctaggg ggaggctaac tgaaacacgg aaggagacaa taccggaagg aacccgcgct
     4921 atgacggcaa taaaaagaca gaataaaacg cacggtgttg ggtcgtttgt tcataaacgc
     4981 ggggttcggt cccagggctg gcactctgtc gataccccac cgagacccca ttggggccaa
     5041 tacgcccgcg tttcttcctt ttccccaccc caccccccaa gttcgggtga aggcccaggg
     5101 ctcgcagcca acgtcggggc ggcaggccct gccatagcct caggttactc atatatactt
     5161 tagattgatt taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat
     5221 aatctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     5281 gaaaagatca aaggatcttc
//
