LOCUS       sequence-90385-e        5994 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses nuclear-targeted human actin, with an mCherry expression 
            reporter after a P2A protease cleavage site, on a CMV promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5994)
  AUTHORS   Belin BJ, Lee T, Mullins RD
  TITLE     DNA damage induces nuclear actin filament assembly by formin-2 and 
            Spire-1/2 that promotes efficient DNA repair.
  JOURNAL   Elife. 2015 Aug 19;4. doi: 10.7554/eLife.07735.
  PUBMED    26287480
REFERENCE   2  (bases 1 to 5994)
  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: "Elife."; 
            date: "2015-08-19"; volume: "4. doi"; pages: " 10.7554/eLife.07735"
COMMENT     Sequence Label: pmCherry-C1 actin-3XNLS P2A mCherry
FEATURES             Location/Qualifiers
     source          1..5994
                     /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"
     CDS             1738..1758
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1762..1782
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1813..1869
                     /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"
     regulatory      1879..1888
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1885..2592
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(2030..2048)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2298..2317
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    2593..2658
                     /label=MCS
                     /note="multiple cloning site"
     polyA_signal    2782..2903
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2819..2838)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2873..2892
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(2910..3365)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3047..3068)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3259..3278
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3392..3496
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3498..3855
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3498..3689
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3521..3541)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3706..3841
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             3890..4684
                     /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             3890..4684
                     /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(3944..3963)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4554..4573
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(4872..4891)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4916..4963
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      5292..5880
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5781..5800
                     /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 gctagcgatg
      601 gatgatgata tcgccgcgct cgtcgtcgac aacggctccg gcatgtgcaa ggccggcttc
      661 gcgggcgacg atgccccccg ggccgtcttc ccctccatcg tggggcgccc caggcaccag
      721 ggcgtgatgg tgggcatggg tcagaaggat tcctatgtgg gcgacgaggc ccagagcaag
      781 agaggcatcc tcaccctgaa gtaccccatc gagcacggca tcgtcaccaa ctgggacgac
      841 atggagaaaa tctggcacca caccttctac aatgagctgc gtgtggctcc cgaggagcac
      901 cccgtgctgc tgaccgaggc ccccctgaac cccaaggcca accgcgagaa gatgacccag
      961 atcatgtttg agaccttcaa caccccagcc atgtacgttg ctatccaggc tgtgctatcc
     1021 ctgtacgcct ctggccgtac cactggcatc gtgatggact ccggtgacgg ggtcacccac
     1081 actgtgccca tctacgaggg gtatgccctc ccccatgcca tcctgcgtct ggacctggct
     1141 ggccgggacc tgactgacta cctcatgaag atcctcaccg agcgcggcta cagcttcacc
     1201 accacggccg agcgggaaat cgtgcgtgac attaaggaga agctgtgcta cgtcgccctg
     1261 gacttcgagc aagagatggc cacggctgct tccagctcct ccctggagaa gagctacgag
     1321 ctgcctgacg gccaggtcat caccattggc aatgagcggt tccgctgccc tgaggcactc
     1381 ttccagcctt ccttcctggg catggagtcc tgtggcatcc acgaaactac cttcaactcc
     1441 atcatgaagt gtgacgtgga catccgcaaa gacctgtacg ccaacacagt gctgtctggc
     1501 ggcaccacca tgtaccctgg cattgccgac aggatgcaga aggagatcac tgccctggca
     1561 cccagcacaa tgaagatcaa gatcattgct cctcctgagc gcaagtactc cgtgtggatc
     1621 ggcggctcca tcctggcctc gctgtccacc ttccagcaga tgtggatcag caagcaggag
     1681 tatgacgagt ccggcccctc catcgtccac cgcaaatgct tcctaccggt aatggatcca
     1741 aaaaagaaga gaaaggtaga tccaaaaaag aagagaaagg tagatccaaa aaagaagaga
     1801 aaggcggccg cagccacgaa cttctctctg ttaaagcaag caggagacgt ggaagaaaac
     1861 cccggtccta aaccggtcgc caccatggtg agcaagggcg aggaggataa catggccatc
     1921 atcaaggagt tcatgcgctt caaggtgcac atggagggct ccgtgaacgg ccacgagttc
     1981 gagatcgagg gcgagggcga gggccgcccc tacgagggca cccagaccgc caagctgaag
     2041 gtgaccaagg gtggccccct gcccttcgcc tgggacatcc tgtcccctca gttcatgtac
     2101 ggctccaagg cctacgtgaa gcaccccgcc gacatccccg actacttgaa gctgtccttc
     2161 cccgagggct tcaagtggga gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg
     2221 acccaggact cctccctgca ggacggcgag ttcatctaca aggtgaagct gcgcggcacc
     2281 aacttcccct ccgacggccc cgtaatgcag aagaagacca tgggctggga ggcctcctcc
     2341 gagcggatgt accccgagga cggcgccctg aagggcgaga tcaagcagag gctgaagctg
     2401 aaggacggcg gccactacga cgctgaggtc aagaccacct acaaggccaa gaagcccgtg
     2461 cagctgcccg gcgcctacaa cgtcaacatc aagttggaca tcacctccca caacgaggac
     2521 tacaccatcg tggaacagta cgaacgcgcc gagggccgcc actccaccgg cggcatggac
     2581 gagctgtaca agtccggact cagatctcga gctcaagctt cgaattctgc agtcgacggt
     2641 accgcgggcc cgggatccac cggatctaga taactgatca taatcagcca taccacattt
     2701 gtagaggttt tacttgcttt aaaaaacctc ccacacctcc ccctgaacct gaaacataaa
     2761 atgaatgcaa ttgttgttgt taacttgttt attgcagctt ataatggtta caaataaagc
     2821 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     2881 tccaaactca tcaatgtatc ttaacgcgta aattgtaagc gttaatattt tgttaaaatt
     2941 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat
     3001 cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa
     3061 gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg
     3121 cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa
     3181 agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc
     3241 gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag
     3301 tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg
     3361 cgcgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     3421 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     3481 ttgaaaaagg aagagtcctg aggcggaaag aaccagctgt ggaatgtgtg tcagttaggg
     3541 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     3601 tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
     3661 catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact
     3721 ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag
     3781 gccgaggccg cctcggcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc
     3841 ctaggctttt gcaaagatcg atcaagagac aggatgagga tcgtttcgca tgattgaaca
     3901 agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg gctatgactg
     3961 ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg
     4021 cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc aagacgaggc
     4081 agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt
     4141 cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg atctcctgtc
     4201 atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc ggcggctgca
     4261 tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca tcgagcgagc
     4321 acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag agcatcaggg
     4381 gctcgcgcca gccgaactgt tcgccaggct caaggcgagc atgcccgacg gcgaggatct
     4441 cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc
     4501 tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc
     4561 tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta
     4621 cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg acgagttctt
     4681 ctgagcggga ctctggggtt cgaaatgacc gaccaagcga cgcccaacct gccatcacga
     4741 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac
     4801 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccctagg
     4861 gggaggctaa ctgaaacacg gaaggagaca ataccggaag gaacccgcgc tatgacggca
     4921 ataaaaagac agaataaaac gcacggtgtt gggtcgtttg ttcataaacg cggggttcgg
     4981 tcccagggct ggcactctgt cgatacccca ccgagacccc attggggcca atacgcccgc
     5041 gtttcttcct tttccccacc ccacccccca agttcgggtg aaggcccagg gctcgcagcc
     5101 aacgtcgggg cggcaggccc tgccatagcc tcaggttact catatatact ttagattgat
     5161 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     5221 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     5281 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     5341 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     5401 gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta
     5461 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     5521 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     5581 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     5641 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     5701 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     5761 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     5821 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     5881 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     5941 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgccat gcat
//
