LOCUS       sequence-470404-        3788 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of mCardinal and extra insert, for MuSIC 
            barcode construction.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3788)
  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 3788)
  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-mCardinal
FEATURES             Location/Qualifiers
     source          1..3788
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    416..537
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      591..1179
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1080..1099
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        1354..1657
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1658..1861
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1812..1832
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             1888..1944
                     /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"
     CDS             1954..2007
                     /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"
     regulatory      2017..2026
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2023..2757
                     /codon_start=1
                     /product="far-red fluorescent protein derived from 
                     mNeptune, with further red-shifted spectra"
                     /label=mCardinal
                     /translation="MVSKGEELIKENMHMKLYMEGTVNNHHFKCTTEGEGKPYEGTQTQ
                     RIKVVEGGPLPFAFDILATCFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGG
                     VLTVTQDTSLQDGCLIYNVKLRGVNFPSNGPVMQKKTLGWEATTETLYPADGGLEGRCD
                     MALKLVGGGHLHCNLKTTYRSKKPAKNLKMPGVYFVDRRLERIKEADNETYVEQHEVAV
                     ARYCDLPSKLGHKLNGMDELYK"
     polyA_signal    2792..2913
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(2920..3375)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3057..3078)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3269..3288
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3401..3503
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             join(3504..3788,1..375)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(3570..3589)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
ORIGIN
        1 catgagcaaa ctgaaacgtt ttcatcgctc tggagtgaat accacgacga tttccggcag
       61 tttctacaca tatattcgca agatgtggcg tgttacggtg aaaacctggc ctatttccct
      121 aaagggttta ttgagaatat gtttttcgta tcagccaatc cctgggtgag tttcaccagt
      181 tttgatttaa acgtggccaa tatggacaac ttcttcgccc ccgttttcac catgggcaaa
      241 tattatacgc aaggcgacaa ggtgctgatg ccgctggcga ttcaggttca tcatgccgtt
      301 tgtgatggct tccatgtcgg cagaatgctt aatgaattac aacagtactg cgatgagtgg
      361 cagggcgggg cgtaatcaaa catgagaatt ggtcgcgcga gcaattcaga catgataaga
      421 tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg ctttatttgt
      481 gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa acaagttggc
      541 atttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
      601 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
      661 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
      721 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
      781 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
      841 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
      901 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
      961 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     1021 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     1081 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     1141 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     1201 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     1261 gattctgtgg ataaccgtat taccgccatg cattagttat taatagtaat caattacggg
     1321 gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc
     1381 gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat
     1441 agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc
     1501 ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga
     1561 cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg
     1621 gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacat
     1681 caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt
     1741 caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactc
     1801 cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagagc
     1861 tggtttagtg aagaagacgg ctccggcgcc acaaacttct ctctgctaaa gcaagcaggt
     1921 gatgttgaag aaaaccccgg gcctggcagc ggcgagggca ggggaagtct acttacatgc
     1981 ggggacgtgg aggaaaatcc cggcccagga tccgccgcca ccatggtgag caagggcgag
     2041 gagctgatca aggagaacat gcacatgaag ctgtacatgg aaggcaccgt gaacaaccac
     2101 cacttcaagt gcaccaccga aggggagggc aagccctacg agggcaccca gacccagagg
     2161 attaaggtgg tggagggagg ccccctgccg ttcgcattcg acatcctggc cacctgcttt
     2221 atgtacggga gcaagacctt catcaaccac acccagggca tccccgattt ctttaagcag
     2281 tccttccctg agggcttcac atgggagaga gtcaccacat acgaggacgg gggcgtgctt
     2341 accgttaccc aggacaccag cctccaggac ggctgcttga tctacaacgt caagctcaga
     2401 ggggtgaact tcccatccaa cggccctgtg atgcagaaga aaacactcgg ctgggaggcc
     2461 accaccgaga ctctgtaccc cgctgacggc ggcctggaag gcagatgcga catggccctg
     2521 aagctcgtgg gcgggggcca cctgcactgc aaccttaaga ccacatacag atccaagaaa
     2581 cccgctaaga acctcaagat gcccggcgtc tactttgtgg accgcagact ggaaagaatc
     2641 aaggaggccg acaatgagac atacgtcgag cagcacgagg tggctgtggc cagatactgc
     2701 gacctcccta gcaaactggg gcacaaactt aatggcatgg acgagctgta caagtaagtt
     2761 cacgtattca tgtcttctga aacataaaat gaacttgttt attgcagctt ataatggtta
     2821 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     2881 ttgtggtttg tccaaactca tcaatgtatc ttaaggcgta aattgtaagc gttaatattt
     2941 tgttaaaatt cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa
     3001 tcggcaaaat cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag
     3061 tttggaacaa gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg
     3121 tctatcaggg cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga
     3181 ggtgccgtaa agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg
     3241 gaaagccggc gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg
     3301 cgctggcaag tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc
     3361 cgctacaggg cgcgtcagta cttaacggaa gatcacttcg tgatcggcac gtaagaggtt
     3421 ccaactttca ccataatgaa ataagatcac taccgggcgt attttttgag ttatcgagat
     3481 tttcaggagc taaggaagct aaaatggaga aaaaaatcac tggatatacc accgttgata
     3541 tatcccaatg gcatcgtaaa gaacattttg aggcatttca gtcagttgct caatgtacct
     3601 ataaccagac cgttcagctg gatattacgg cctttttaaa gaccgtaaag aaaaataagc
     3661 acaagtttta tccggccttt attcacattc ttgcccgcct gatgaatgct catccggaat
     3721 tccgtatggc aatgaaagac ggtgagctgg tgatatggga tagtgttcac ccttgttaca
     3781 ccgttttc
//
