LOCUS       sequence-468966-        4409 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of miRFP670, for MuSIC barcode construction.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4409)
  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 4409)
  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-miRFP670
FEATURES             Location/Qualifiers
     source          1..4409
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(97..891)
                     /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(97..891)
                     /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(208..227)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     818..837
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(926..1122)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        complement(934..1144)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      940..1075
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1168..1188
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      1232..1687
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1319..1338)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1529..1550
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     polyA_signal    1694..1815
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1705..1724)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1759..1778
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(1894..2838)
                     /codon_start=1
                     /product="bright monomeric near-infrared fluorescent 
                     protein engineered from a bacterial phytochrome, with an 
                     emission maximum of 670 nm (Shcherbakova et al., 2016)"
                     /label=miRFP670
                     /label=ORF frame 1
                     /translation="MVAGHASGSPAFGTASHSNCEHEEIHLAGSIQPHGALLVVSEHDH
                     RVIQASANAAEFLNLGSVLGVPLAEIDGDLLIKILPHLDPTAEGMPVAVRCRIGNPSTE
                     YCGLMHRPPEGGLIIELERAGPSIDLSGTLAPALERIRTAGSLRALCDDTVLLFQQCTG
                     YDRVMVYRFDEQGHGLVFSECHVPGLESYFGNRYPSSTVPQMARQLYVRQRVRVLVDVT
                     YQPVPLEPRLSPLTGRDLDMSGCFLRSMSPCHLQFLKDMGVRATLAVSLVVGGKLWGLV
                     VCHHYLPRFIRFELRAICKRLAERIATRITALES"
     regulatory      2835..2844
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(2892..3095)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2921..2941)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        3096..3399
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     rep_origin      complement(3580..4168)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3660..3679)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     polyA_signal    4227..4274
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     4299..4318
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
ORIGIN
        1 agcccaacct ttcatagaag gcggcggtgg aatcgaaatc tcgtgatggc aggttgggcg
       61 tcgcttggtc ggtcatttcg aaccccagag tcccgctcag aagaactcgt caagaaggcg
      121 atagaaggcg atgcgctgcg aatcgggagc ggcgataccg taaagcacga ggaagcggtc
      181 agcccattcg ccgccaagct cttcagcaat atcacgggta gccaacgcta tgtcctgata
      241 gcggtccgcc acacccagcc ggccacagtc gatgaatcca gaaaagcggc cattttccac
      301 catgatattc ggcaagcagg catcgccatg ggtcacgacg agatcctcgc cgtcgggcat
      361 gctcgccttg agcctggcga acagttcggc tggcgcgagc ccctgatgct cttcgtccag
      421 atcatcctga tcgacaagac cggcttccat ccgagtacgt gctcgctcga tgcgatgttt
      481 cgcttggtgg tcgaatgggc aggtagccgg atcaagcgta tgcagccgcc gcattgcatc
      541 agccatgatg gatactttct cggcaggagc aaggtgagat gacaggagat cctgccccgg
      601 cacttcgccc aatagcagcc agtcccttcc cgcttcagtg acaacgtcga gcacagctgc
      661 gcaaggaacg cccgtcgtgg ccagccacga tagccgcgct gcctcgtctt gcagttcatt
      721 cagggcaccg gacaggtcgg tcttgacaaa aagaaccggg cgcccctgcg ctgacagccg
      781 gaacacggcg gcatcagagc agccgattgt ctgttgtgcc cagtcatagc cgaatagcct
      841 ctccacccaa gcggccggag aacctgcgtg caatccatct tgttcaatca tgcgaaacga
      901 tcctcatcct gtctcttgat cgatctttgc aaaagcctag gcctccaaaa aagcctcctc
      961 actacttctg gaatagctca gaggccgagg cggcctcggc ctctgcataa ataaaaaaaa
     1021 ttagtcagcc atggggcgga gaatgggcgg aactgggcgg agttaggggc gggatgggcg
     1081 gagttagggg cgggactatg gttgctgact aattgagatg catgctttgc atacttctgc
     1141 ctgctgggga gcctggggac tttccacacc ctaactgaca cacattccac agctggttct
     1201 ttccgcctca ggactcttcc tttaagtact gacgcgccct gtagcggcgc attaagcgcg
     1261 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
     1321 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     1381 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     1441 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     1501 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     1561 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     1621 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgctt
     1681 acaatttacg ccttaagata cattgatgag tttggacaaa ccacaactag aatgcagtga
     1741 aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc
     1801 tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcagaa ttcattatct
     1861 agaattcgtc ttcaattatg aagacatgga gccgctctca agcgcggtga tccgcgtcgc
     1921 gatcctttcg gcgagccgtt tgcagatcgc ccgcagctcg aaacggatga agcgcggcag
     1981 ataatggtga cagacaacca ggccccacag cttgccgccg accaccagcg acaccgccag
     2041 ggtggcgcgc acgcccatgt ccttcaggaa ctgcagatgg cacggcgaca tcgagcgcag
     2101 gaagcagccc gacatgtcga gatcgcgccc ggtcagcggc gacagccgcg gctccagcgg
     2161 caccggctga taggtgacgt cgaccagcac gcggacgcgc tgccgcacgt acagctgccg
     2221 cgccatctgc gggacagtcg acgacggata gcggttgccg aaataggatt cgagcccagg
     2281 cacatggcac tcggagaata ccaggccgtg gccttgctca tcgaaacgat acaccatcac
     2341 ccggtcgtag ccggtgcact gctgaaacag cagcacggtg tcatcgcaca gcgcgcgcag
     2401 tgaacccgcc gtgcggatcc gctccagcgc cggcgccagc gtgcctgaca gatcgatcga
     2461 cgggccggca cgttcgagtt cgatgatcag cccgccttcc ggaggccgat gcatcagacc
     2521 gcagtactcc gtagagggat tgccgatccg gcagcgcacc gcgaccggca tgccttcggc
     2581 ggtgggatcg agatgcggca ggatcttgat caacagatcg ccgtcgatct cggcgagcgg
     2641 aacgccgagt acgcttccga gattcagaaa ttccgcggcg ttggcgctgg cctggatgac
     2701 gcgatgatca tgttcgctga cgaccagaag cgcgccatgc ggctggatcg agccggcgag
     2761 gtggatctct tcatgttcgc aattcgaatg agaggcggtc ccgaatgcgg ggctgccaga
     2821 ggcatgacct gctaccatgg tggcggcgga tccatttcga taagccagta agcacgaggc
     2881 tgatcagcgg tagctctgct tatatagacc tcccaccgta cacgcctacc gcccatttgc
     2941 gtcaatgggg cggagttgtt acgacatttt ggaaagtccc gttgattttg gtgccaaaac
     3001 aaactcccat tgacgtcaat ggggtggaga cttggaaatc cccgtgagtc aaaccgctat
     3061 ccacgcccat tgatgtactg ccaaaaccgc atcaccatgg taatagcgat gactaatacg
     3121 tagatgtact gccaagtagg aaagtcccat aaggtcatgt actgggcata atgccaggcg
     3181 ggccatttac cgtcattgac gtcaataggg ggcgtacttg gcatatgata cacttgatgt
     3241 actgccaagt gggcagttta ccgtaaatac tccacccatt gacgtcaatg gaaagtccct
     3301 attggcgtta ctatgggaac atacgtcatt attgacgtca atgggcgggg gtcgttgggc
     3361 ggtcagccag gcgggccatt taccgtaagt tatgtaacgg ctagccggaa ctccatatat
     3421 gggctatgaa ctaatgaccc cgtaattgat tactattaat aactaatgca tggcggtaat
     3481 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     3541 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     3601 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     3661 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     3721 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     3781 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     3841 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     3901 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     3961 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     4021 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     4081 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     4141 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     4201 cgctcagtgg aacgaaccgc gtttatgaac aaacgaccca acaccgtgcg ttttattctg
     4261 tctttttatt gccgtcatag cgcgggttcc ttccggtatt gtctccttcc gtgtttcagt
     4321 tagcctcccc ctagggtggg cgaagaactc cagcatgaga tccccgcgct ggaggatcat
     4381 ccagccggcg tcccggaaaa cgattccga
//
