LOCUS       sequence-457667-        3752 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of mKate2 and extra insert, for MuSIC barcode 
            construction.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3752)
  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 3752)
  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: pM-mKate2
FEATURES             Location/Qualifiers
     source          1..3752
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(50..748)
                     /codon_start=1
                     /product="monomeric far-red fluorescent protein (Shcherbo 
                     et al., 2009)"
                     /label=mKate2
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGHR"
     regulatory      745..754
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             complement(764..817)
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             complement(827..883)
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     promoter        complement(910..1113)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(939..959)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        1114..1417
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(1592..2180)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1672..1691)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    2234..2355
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             complement(2396..3055)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     2970..2989
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(3056..3158)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     rep_origin      3184..3639
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3271..3290)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3481..3502
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    join(3646..3752,1..15)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
ORIGIN
        1 tgcaataaac aagttcattt tatgtttcag aagacatgaa tggcaactct tatctgtgcc
       61 ccagtttgct agggaggtcg cagtatctgg ccacagccac ctcgtgctgc tcgacgtaag
      121 tctctttgtc ggcctccttg attctttcca gccttctgtc cacatagtag acgccgggca
      181 tcttgaggtt cttagcgggt ttcttggatc tgtatgtggt ctttaagttg cagatcaggt
      241 ggcccccgcc cacgagcttc agggccatgt cggctctgcc ttccaggccg ccgtcagcgg
      301 ggtacagtgt ctcggtggag gcctcccagc cgagtgtttt cttctgcatc acagggccgt
      361 tggatgggaa gttcacccct ctgatcttga cgttgtagat gaggcagccg tcctggaggc
      421 tggtgtcctg ggtagcggtc agcacgcccc catcttcgta tgtggtgact ctctcccatg
      481 tgaagccctc ggggaaggac tgcttaaaga agtcggggat gccctgggtg tggttgatga
      541 aggttttgct gccgtacatg aagctggtag ccaggatgtc gaaggcgaag gggagagggc
      601 cgccctcgac cgccttgatt ctcatggtct gggtgccctc gtagggcttg ccttcgccct
      661 cggatgtgca cttgaagtgg tggttgttca cggtgccctc catgtacagc ttcatgtgca
      721 tgttctcctt aatcagctcg ctcaccatgg tggcggcgga tcctgggccg ggattttcct
      781 ccacgtcccc gcatgtaagt agacttcccc tgccctcgcc gctgccaggc ccggggtttt
      841 cttcaacatc acctgcttgc tttagcagag agaagtttgt ggcgccggag ccgtcttctt
      901 cactaaacca gctctgctta tatagacctc ccaccgtaca cgcctaccgc ccatttgcgt
      961 caatggggcg gagttgttac gacattttgg aaagtcccgt tgattttggt gccaaaacaa
     1021 actcccattg acgtcaatgg ggtggagact tggaaatccc cgtgagtcaa accgctatcc
     1081 acgcccattg atgtactgcc aaaaccgcat caccatggta atagcgatga ctaatacgta
     1141 gatgtactgc caagtaggaa agtcccataa ggtcatgtac tgggcataat gccaggcggg
     1201 ccatttaccg tcattgacgt caataggggg cgtacttggc atatgataca cttgatgtac
     1261 tgccaagtgg gcagtttacc gtaaatactc cacccattga cgtcaatgga aagtccctat
     1321 tggcgttact atgggaacat acgtcattat tgacgtcaat gggcgggggt cgttgggcgg
     1381 tcagccaggc gggccattta ccgtaagtta tgtaacgcgg aactccatat atgggctatg
     1441 aactaatgac cccgtaattg attactatta ataactaatg catggcggta atacggttat
     1501 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
     1561 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
     1621 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
     1681 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
     1741 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
     1801 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
     1861 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
     1921 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
     1981 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
     2041 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
     2101 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
     2161 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
     2221 ggaacgaaat gccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     2281 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     2341 catcaatgta tcttatcatg tctgaattgc tcgcgcgacc aattctcatg tttgattacg
     2401 ccccgccctg ccactcatcg cagtactgtt gtaattcatt aagcattctg ccgacatgga
     2461 agccatcaca aacggcatga tgaacctgaa tcgccagcgg catcagcacc ttgtcgcctt
     2521 gcgtataata tttgcccatg gtgaaaacgg gggcgaagaa gttgtccata ttggccacgt
     2581 ttaaatcaaa actggtgaaa ctcacccagg gattggctga tacgaaaaac atattctcaa
     2641 taaacccttt agggaaatag gccaggtttt caccgtaaca cgccacatct tgcgaatata
     2701 tgtgtagaaa ctgccggaaa tcgtcgtggt attcactcca gagcgatgaa aacgtttcag
     2761 tttgctcatg gaaaacggtg taacaagggt gaacactatc ccatatcacc agctcaccgt
     2821 ctttcattgc catacggaat tccggatgag cattcatcag gcgggcaaga atgtgaataa
     2881 aggccggata aaacttgtgc ttatttttct ttacggtctt taaaaaggcc gtaatatcca
     2941 gctgaacggt ctggttatag gtacattgag caactgactg aaatgcctca aaatgttctt
     3001 tacgatgcca ttgggatata tcaacggtgg tatatccagt gatttttttc tccattttag
     3061 cttccttagc tcctgaaaat ctcgataact caaaaaatac gcccggtagt gatcttattt
     3121 cattatggtg aaagttggaa cctcttacgt gccgatcacg aagtgatctt ccgttaagta
     3181 ctgacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     3241 ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg
     3301 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     3361 ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt tcacgtagtg
     3421 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     3481 gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat tcttttgatt
     3541 tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat
     3601 ttaacgcgaa ttttaacaaa atattaacgc ttacaattta cgccttaaga tacattgatg
     3661 agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg
     3721 atgctattgc tttatttgta accattataa gc
//
