LOCUS       sequence-452465-        4181 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of mT-Sapphire, for MuSIC barcode construction.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4181)
  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 4181)
  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-mT-Sapphire
FEATURES             Location/Qualifiers
     source          1..4181
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(98..892)
                     /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(98..892)
                     /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(209..228)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     819..838
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(927..1123)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        complement(935..1145)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      941..1076
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1169..1189
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      1233..1688
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1320..1339)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1530..1551
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    1695..1816
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1706..1725)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1760..1779
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(join(1895..2605,2606..2608,2609..2611))
                     /codon_start=1
                     /product="strongly enhanced green fluorescent protein 
                     (Kremers et al., 2007)"
                     /label=SGFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFSYGVMVFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYIMADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSIQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1937..1958)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2286..2305
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2545..2566
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      2608..2617
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(2665..2868)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2694..2714)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2869..3172
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(3353..3941)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3433..3452)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    4000..4047
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     4072..4091
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
ORIGIN
        1 aagcccaacc tttcatagaa ggcggcggtg gaatcgaaat ctcgtgatgg caggttgggc
       61 gtcgcttggt cggtcatttc gaaccccaga gtcccgctca gaagaactcg tcaagaaggc
      121 gatagaaggc gatgcgctgc gaatcgggag cggcgatacc gtaaagcacg aggaagcggt
      181 cagcccattc gccgccaagc tcttcagcaa tatcacgggt agccaacgct atgtcctgat
      241 agcggtccgc cacacccagc cggccacagt cgatgaatcc agaaaagcgg ccattttcca
      301 ccatgatatt cggcaagcag gcatcgccat gggtcacgac gagatcctcg ccgtcgggca
      361 tgctcgcctt gagcctggcg aacagttcgg ctggcgcgag cccctgatgc tcttcgtcca
      421 gatcatcctg atcgacaaga ccggcttcca tccgagtacg tgctcgctcg atgcgatgtt
      481 tcgcttggtg gtcgaatggg caggtagccg gatcaagcgt atgcagccgc cgcattgcat
      541 cagccatgat ggatactttc tcggcaggag caaggtgaga tgacaggaga tcctgccccg
      601 gcacttcgcc caatagcagc cagtcccttc ccgcttcagt gacaacgtcg agcacagctg
      661 cgcaaggaac gcccgtcgtg gccagccacg atagccgcgc tgcctcgtct tgcagttcat
      721 tcagggcacc ggacaggtcg gtcttgacaa aaagaaccgg gcgcccctgc gctgacagcc
      781 ggaacacggc ggcatcagag cagccgattg tctgttgtgc ccagtcatag ccgaatagcc
      841 tctccaccca agcggccgga gaacctgcgt gcaatccatc ttgttcaatc atgcgaaacg
      901 atcctcatcc tgtctcttga tcgatctttg caaaagccta ggcctccaaa aaagcctcct
      961 cactacttct ggaatagctc agaggccgag gcggcctcgg cctctgcata aataaaaaaa
     1021 attagtcagc catggggcgg agaatgggcg gaactgggcg gagttagggg cgggatgggc
     1081 ggagttaggg gcgggactat ggttgctgac taattgagat gcatgctttg catacttctg
     1141 cctgctgggg agcctgggga ctttccacac cctaactgac acacattcca cagctggttc
     1201 tttccgcctc aggactcttc ctttaagtac tgacgcgccc tgtagcggcg cattaagcgc
     1261 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     1321 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     1381 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     1441 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     1501 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     1561 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg
     1621 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgct
     1681 tacaatttac gccttaagat acattgatga gtttggacaa accacaacta gaatgcagtg
     1741 aaaaaaatgc tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccattataag
     1801 ctgcaataaa caagttaaca acaacaattg cattcatttt atgtttcaga attcattatc
     1861 tagaattcgt cttcaattat gaagacatgg agcccttgta cagctcgtcc atgccgagag
     1921 tgatcccggc ggcggtcacg aactccagca ggaccatgtg atcgcgcttc tcgttggggt
     1981 ctttgctcag cttggactgg atgctcaggt agtggttgtc gggcagcagc acggggccgt
     2041 cgccgatggg ggtgttctgc tggtagtggt cggcgagctg cacgccgccg tcctcgatgt
     2101 tgtggcggat cttgaagttg gccttgatgc cgttcttctg cttgtcggcc atgatataga
     2161 cgttgtggct gttgaagttg tactccagct tgtgccccag gatgttgccg tcctccttga
     2221 agtcgatgcc cttcagctcg atgcggttca ccagggtgtc gccctcgaac ttcacctcgg
     2281 cgcgggtctt gtagttgccg tcgtccttga agaagatggt gcgctcctgg acgtagcctt
     2341 cgggcatggc ggacttgaag aagtcgtgct gcttcatgtg gtcggggtag cgggcgaaca
     2401 ccatcacgcc gtaggagaag gtggtcacga gggtgggcca gggcacgggc agcttgccgg
     2461 tggtgcagat gaacttcagg gtcagcttgc cgtaggtggc atcgccctcg ccctcgccgg
     2521 acacgctgaa cttgtggccg tttacgtcgc cgtccagctc gaccaggatg ggcaccaccc
     2581 cggtgaacag ctcctcgccc ttgctcacca tggtggcggc ggatccattt cgataagcca
     2641 gtaagcacga ggctgatcag cggtagctct gcttatatag acctcccacc gtacacgcct
     2701 accgcccatt tgcgtcaatg gggcggagtt gttacgacat tttggaaagt cccgttgatt
     2761 ttggtgccaa aacaaactcc cattgacgtc aatggggtgg agacttggaa atccccgtga
     2821 gtcaaaccgc tatccacgcc cattgatgta ctgccaaaac cgcatcacca tggtaatagc
     2881 gatgactaat acgtagatgt actgccaagt aggaaagtcc cataaggtca tgtactgggc
     2941 ataatgccag gcgggccatt taccgtcatt gacgtcaata gggggcgtac ttggcatatg
     3001 atacacttga tgtactgcca agtgggcagt ttaccgtaaa tactccaccc attgacgtca
     3061 atggaaagtc cctattggcg ttactatggg aacatacgtc attattgacg tcaatgggcg
     3121 ggggtcgttg ggcggtcagc caggcgggcc atttaccgta agttatgtaa cggctagccg
     3181 gaactccata tatgggctat gaactaatga ccccgtaatt gattactatt aataactaat
     3241 gcatggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     3301 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     3361 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     3421 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     3481 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     3541 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     3601 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     3661 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     3721 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     3781 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     3841 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     3901 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     3961 ctacggggtc tgacgctcag tggaacgaac cgcgtttatg aacaaacgac ccaacaccgt
     4021 gcgttttatt ctgtcttttt attgccgtca tagcgcgggt tccttccggt attgtctcct
     4081 tccgtgtttc agttagcctc cccctagggt gggcgaagaa ctccagcatg agatccccgc
     4141 gctggaggat catccagccg gcgtcccgga aaacgattcc g
//
