LOCUS       sequence-383393-        6066 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the fusion protein of miRFP670-2-Actin-C-18 in mammalian 
            cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6066)
  AUTHORS   Zhang H, Papadaki S, Sun X, Wang X, Drobizhev M, Yao L, Rehbock M, 
            Koster RW, Wu L, Namikawa K, Piatkevich KD
  TITLE     Quantitative assessment of near-infrared fluorescent proteins.
  JOURNAL   Nat Methods. 2023 Sep 4. doi: 10.1038/s41592-023-01975-z.
  PUBMED    37666982
REFERENCE   2  (bases 1 to 6066)
  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"; journalName: "Nat 
            Methods. 2023 Sep 4. doi: 10.1038/s41592-023-01975-z."
COMMENT     Sequence Label: pmiRFP670-2-Actin-C-18
FEATURES             Location/Qualifiers
     source          1..6066
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        365..568
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     519..539
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      607..616
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             613..1545
                     /codon_start=1
                     /product="near-infrared fluorescent protein with an 
                     emission peak at 670 nm, engineered from a bacterial 
                     phytochrome (Shcherbakova and Verkhusha, 2013)"
                     /label=iRFP670
                     /note="derived from Rhodopseudomonas palustris RpBphP6"
                     /translation="MARKVDLTSCDREPIHIPGSIQPCGCLLACDAQAVRITRITENAG
                     AFFGRETPRVGELLADYFGETEAHALRNALAQSSDPKRPALIFGWRDGLTGRTFDISLH
                     RHDGTSIIEFEPAAAEQADNPLRLTRQIIARTKELKSLEEMAARVPRYLQAMLGYHRVM
                     LYRFADDGSGMVIGEAKRSDLESFLGQHFPASLVPQQARLLYLKNAIRVVSDSRGISSR
                     IVPEHDASGAALDLSFAHLRSISPCHLEFLRNMGVSASMSLSIIIDGTLWGLIICHHYE
                     PRAVPMAQRVAAKRFAERLSTHFTAAHHQR"
     polyA_signal    2854..2975
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2891..2910)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2945..2964
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2982..3437)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3119..3140)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3331..3350
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3464..3568
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3570..3927
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3570..3761
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3593..3613)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3778..3913
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3962..4756
                     /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             3962..4756
                     /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(4016..4035)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4626..4645
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4944..4963)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4988..5035
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5364..5952
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5853..5872
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      421 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      481 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      541 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      601 ccggtcgcca ccatggcgcg taaggtcgat ctcacctcct gcgatcgcga gccgatccac
      661 atccccggca gcattcagcc gtgcggctgc ctgctagcct gcgacgcgca ggcggtgcgg
      721 atcacgcgca ttacggaaaa tgccggcgcg ttctttggac gcgaaactcc gcgggtcggt
      781 gagctactcg ccgattactt cggcgagacc gaagcccatg cgctgcgcaa cgcactggcg
      841 cagtcctccg atccaaagcg accggcgctg atcttcggtt ggcgcgacgg cctgaccggc
      901 cgcaccttcg acatctcact gcatcgccat gacggtacat cgatcatcga gttcgagcct
      961 gcggcggccg aacaggccga caatccgctg cggctgacgc ggcagatcat cgcgcgcacc
     1021 aaagaactga agtcgctcga agagatggcc gcacgggtgc cgcgctatct gcaggcgatg
     1081 ctcggctatc accgcgtgat gttgtaccgc ttcgcggacg acggctccgg gatggtgatc
     1141 ggcgaggcga agcgcagcga cctcgagagc tttctcggtc agcactttcc ggcgtcgctg
     1201 gtcccgcagc aggcgcggct actgtacttg aagaacgcga tccgcgtggt ctcggattcg
     1261 cgcggcatca gcagccggat cgtgcccgag cacgacgcct ccggcgccgc gctcgatctg
     1321 tcgttcgcgc acctgcgcag catctcgccc tgccatctcg aatttctgcg gaacatgggc
     1381 gtcagcgcct cgatgtcgct gtcgatcatc attgacggca cgctatgggg attgatcatc
     1441 tgtcatcatt acgagccgcg tgccgtgccg atggcgcagc gcgtcgcggc caaaaggttc
     1501 gccgagcgct tatcgacgca cttcaccgcc gcccaccacc aacgctccgg actcagatct
     1561 ggcagcggtg gaggcagcgc atccggcgga agcggaagcg atgatgatat cgccgcgctc
     1621 gtcgtcgaca acggctccgg catgtgcaag gccggcttcg cgggcgacga tgccccccgg
     1681 gccgtcttcc cctccatcgt ggggcgcccc aggcaccagg gcgtgatggt gggcatgggt
     1741 cagaaggatt cctatgtggg cgacgaggcc cagagcaaga gaggcatcct caccctgaag
     1801 taccccatcg agcacggcat cgtcaccaac tgggacgaca tggagaaaat ctggcaccac
     1861 accttctaca atgagctgcg tgtggctccc gaggagcacc ccgtgctgct gaccgaggcc
     1921 cccctgaacc ccaaggccaa ccgcgagaag atgacccaga tcatgtttga gaccttcaac
     1981 accccagcca tgtacgttgc tatccaggct gtgctatccc tgtacgcctc tggccgtacc
     2041 actggcatcg tgatggactc cggtgacggg gtcacccaca ctgtgcccat ctacgagggg
     2101 tatgccctcc cccatgccat cctgcgtctg gacctggctg gccgggacct gactgactac
     2161 ctcatgaaga tcctcaccga gcgcggctac agcttcacca ccacggccga gcgggaaatc
     2221 gtgcgtgaca ttaaggagaa gctgtgctac gtcgccctgg acttcgagca agagatggcc
     2281 acggctgctt ccagctcctc cctggagaag agctacgagc tgcctgacgg ccaggtcatc
     2341 accattggca atgagcggtt ccgctgccct gaggcactct tccagccttc cttcctgggc
     2401 atggagtcct gtggcatcca cgaaactacc ttcaactcca tcatgaagtg tgacgtggac
     2461 atccgcaaag acctgtacgc caacacagtg ctgtctggcg gcaccaccat gtaccctggc
     2521 attgccgaca ggatgcagaa ggagatcact gccctggcac ccagcacaat gaagatcaag
     2581 atcattgctc ctcctgagcg caagtactcc gtgtggatcg gcggctccat cctggcctcg
     2641 ctgtccacct tccagcagat gtggatcagc aagcaggagt atgacgagtc cggcccctcc
     2701 atcgtccacc gcaaatgctt ctagggatcc accggatcta gataactgat cataatcagc
     2761 cataccacat ttgtagaggt tttacttgct ttaaaaaacc tcccacacct ccccctgaac
     2821 ctgaaacata aaatgaatgc aattgttgtt gttaacttgt ttattgcagc ttataatggt
     2881 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     2941 agttgtggtt tgtccaaact catcaatgta tcttaacgcg taaattgtaa gcgttaatat
     3001 tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc aataggccga
     3061 aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga gtgttgttcc
     3121 agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac
     3181 cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt ttttggggtc
     3241 gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta gagcttgacg
     3301 gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag cgggcgctag
     3361 ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc
     3421 gccgctacag ggcgcgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     3481 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     3541 gcttcaataa tattgaaaaa ggaagagtcc tgaggcggaa agaaccagct gtggaatgtg
     3601 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     3661 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
     3721 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
     3781 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
     3841 atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta gtgaggaggc
     3901 ttttttggag gcctaggctt ttgcaaagat cgatcaagag acaggatgag gatcgtttcg
     3961 catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt
     4021 cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc
     4081 agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact
     4141 gcaagacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt
     4201 gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca
     4261 ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat
     4321 gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg
     4381 catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga
     4441 agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcga gcatgcccga
     4501 cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa
     4561 tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga
     4621 catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt
     4681 cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct
     4741 tgacgagttc ttctgagcgg gactctgggg ttcgaaatga ccgaccaagc gacgcccaac
     4801 ctgccatcac gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc
     4861 gttttccggg acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc
     4921 gcccacccta gggggaggct aactgaaaca cggaaggaga caataccgga aggaacccgc
     4981 gctatgacgg caataaaaag acagaataaa acgcacggtg ttgggtcgtt tgttcataaa
     5041 cgcggggttc ggtcccaggg ctggcactct gtcgataccc caccgagacc ccattggggc
     5101 caatacgccc gcgtttcttc cttttcccca ccccaccccc caagttcggg tgaaggccca
     5161 gggctcgcag ccaacgtcgg ggcggcaggc cctgccatag cctcaggtta ctcatatata
     5221 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     5281 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     5341 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     5401 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     5461 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg
     5521 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     5581 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     5641 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     5701 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     5761 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     5821 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     5881 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     5941 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     6001 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     6061 atgcat
//
