LOCUS       sequence-452462-        4175 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of mRuby3, for MuSIC barcode construction.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4175)
  AUTHORS   Lu X, Pritko DJ, Abravanel ME, Huggins JR, Ogunleye O, Biswas T, 
            Ashy KC, Woods SK, Livingston MWT, Blenner MA, Birtwistle MR
  TITLE     Genetically Encoded Fluorescence Barcodes Allow for Single-Cell 
            Analysis via Spectral Flow Cytometry.
  JOURNAL   ACS Synth Biol. 2025 May 16;14(5):1533-1548. doi: 
            10.1021/acssynbio.4c00807. Epub 2025 May 6.
  PUBMED    40326708
REFERENCE   2  (bases 1 to 4175)
  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/acssynbio.4c00807"; journalName: "ACS Synth Biol"; date: 
            "2025-05-16- 16"; volume: "14"; issue: "5"; pages: "1533-1548"
COMMENT     Sequence Label: pR-mRuby3
FEATURES             Location/Qualifiers
     source          1..4175
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     330..349
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(438..634)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        complement(446..656)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      452..587
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     680..700
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      744..1199
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(831..850)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1041..1062
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    1206..1327
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1217..1236)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1271..1290
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(1406..2116)
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby2, with improved brightness and photostability (Bajar 
                     et al., 2016)"
                     /label=mRuby3
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGRPYEGVQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPADIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVTQDTSLEDGELVYNVKVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTD
                     IALKVDGGGHLHCNFVTTYRSKKTVGNIKMPGVHAVDHRLERIEESDNETYVVQREVAV
                     AKYSNLGGGMDELYK"
     regulatory      2113..2122
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(2170..2373)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2199..2219)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2374..2677
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(2858..3446)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2938..2957)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    3505..3552
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     3577..3596
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     CDS             complement(join(3784..4175,1..403))
                     /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             complement(join(3784..4175,1..403))
                     /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(3895..3914)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
ORIGIN
        1 ggtcgaatgg gcaggtagcc ggatcaagcg tatgcagccg ccgcattgca tcagccatga
       61 tggatacttt ctcggcagga gcaaggtgag atgacaggag atcctgcccc ggcacttcgc
      121 ccaatagcag ccagtccctt cccgcttcag tgacaacgtc gagcacagct gcgcaaggaa
      181 cgcccgtcgt ggccagccac gatagccgcg ctgcctcgtc ttgcagttca ttcagggcac
      241 cggacaggtc ggtcttgaca aaaagaaccg ggcgcccctg cgctgacagc cggaacacgg
      301 cggcatcaga gcagccgatt gtctgttgtg cccagtcata gccgaatagc ctctccaccc
      361 aagcggccgg agaacctgcg tgcaatccat cttgttcaat catgcgaaac gatcctcatc
      421 ctgtctcttg atcgatcttt gcaaaagcct aggcctccaa aaaagcctcc tcactacttc
      481 tggaatagct cagaggccga ggcggcctcg gcctctgcat aaataaaaaa aattagtcag
      541 ccatggggcg gagaatgggc ggaactgggc ggagttaggg gcgggatggg cggagttagg
      601 ggcgggacta tggttgctga ctaattgaga tgcatgcttt gcatacttct gcctgctggg
      661 gagcctgggg actttccaca ccctaactga cacacattcc acagctggtt ctttccgcct
      721 caggactctt cctttaagta ctgacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
      781 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc
      841 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
      901 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta
      961 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     1021 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat
     1081 ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa
     1141 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaattta
     1201 cgccttaaga tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg
     1261 ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa
     1321 acaagttaac aacaacaatt gcattcattt tatgtttcag aattccgacc atggaattcg
     1381 tcttcaatta tgaagacatg gagcccttgt acagctcgtc catgccacca ccaaggttgc
     1441 tgtatttggc aactgccact tctctttgca ctacgtaggt ttcattgtca ctctcctcga
     1501 tcctttccag gcggtgatca acggcatgga caccgggcat cttgatgttc ccgacggtct
     1561 tttttgacct gtaagttgtc acgaagttgc agtgcagatg gccaccacca tcaactttca
     1621 gtgcgatgtc agtgtatcct ctcagaccac catctgctgg atacatcatc tctgtattag
     1681 gctcccaacc cttggttttc ttctgcatca cgggaccatt ggagggaaag tttacccctc
     1741 tgaccttgac gttgtagacg agctcgccat cctcaaggct ggtgtcctgc gtgacggtga
     1801 cgactccacc atcttcgtat ctcgtaactc tttcccaagt aaaaccctca ggaaaggact
     1861 gtttaaagaa atcagggatg tcggccgggt acttgataaa ggtacggctg ccatacatga
     1921 acgacgtggc aagaatgtca aaggcaaatg gcaggggtcc tccctcgatg actttgatcc
     1981 tcatggtttg cactccctcg tacggtctgc cttctccttc acctgtgcat ttgaattggt
     2041 ggccgttgac cgaaccttcc atgaccacct tcatacgcat attttccttg atcagctctt
     2101 cgcccttaga caccatggtg gcggcggatc catttcgata agccagtaag cacgaggctg
     2161 atcagcggta gctctgctta tatagacctc ccaccgtaca cgcctaccgc ccatttgcgt
     2221 caatggggcg gagttgttac gacattttgg aaagtcccgt tgattttggt gccaaaacaa
     2281 actcccattg acgtcaatgg ggtggagact tggaaatccc cgtgagtcaa accgctatcc
     2341 acgcccattg atgtactgcc aaaaccgcat caccatggta atagcgatga ctaatacgta
     2401 gatgtactgc caagtaggaa agtcccataa ggtcatgtac tgggcataat gccaggcggg
     2461 ccatttaccg tcattgacgt caataggggg cgtacttggc atatgataca cttgatgtac
     2521 tgccaagtgg gcagtttacc gtaaatactc cacccattga cgtcaatgga aagtccctat
     2581 tggcgttact atgggaacat acgtcattat tgacgtcaat gggcgggggt cgttgggcgg
     2641 tcagccaggc gggccattta ccgtaagtta tgtaacggct agccggaact ccatatatgg
     2701 gctatgaact aatgaccccg taattgatta ctattaataa ctaatgcatg gcggtaatac
     2761 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     2821 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     2881 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     2941 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     3001 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     3061 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     3121 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     3181 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     3241 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     3301 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     3361 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     3421 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     3481 ctcagtggaa cgaaccgcgt ttatgaacaa acgacccaac accgtgcgtt ttattctgtc
     3541 tttttattgc cgtcatagcg cgggttcctt ccggtattgt ctccttccgt gtttcagtta
     3601 gcctccccct agggtgggcg aagaactcca gcatgagatc cccgcgctgg aggatcatcc
     3661 agccggcgtc ccggaaaacg attccgaagc ccaacctttc atagaaggcg gcggtggaat
     3721 cgaaatctcg tgatggcagg ttgggcgtcg cttggtcggt catttcgaac cccagagtcc
     3781 cgctcagaag aactcgtcaa gaaggcgata gaaggcgatg cgctgcgaat cgggagcggc
     3841 gataccgtaa agcacgagga agcggtcagc ccattcgccg ccaagctctt cagcaatatc
     3901 acgggtagcc aacgctatgt cctgatagcg gtccgccaca cccagccggc cacagtcgat
     3961 gaatccagaa aagcggccat tttccaccat gatattcggc aagcaggcat cgccatgggt
     4021 cacgacgaga tcctcgccgt cgggcatgct cgccttgagc ctggcgaaca gttcggctgg
     4081 cgcgagcccc tgatgctctt cgtccagatc atcctgatcg acaagaccgg cttccatccg
     4141 agtacgtgct cgctcgatgc gatgtttcgc ttggt
//
