LOCUS       sequence-399654-        7041 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mtDNA elimination/production of rho-0 cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7041)
  AUTHORS   Spadafora D, Kozhukhar N, Chouljenko VN, Kousoulas KG, Alexeyev MF
  TITLE     Methods for Efficient Elimination of Mitochondrial DNA from Cultured
            Cells.
  JOURNAL   PLoS One. 2016 May 2;11(5):e0154684. doi: 
            10.1371/journal.pone.0154684. eCollection 2016.
  PUBMED    27136098
REFERENCE   2  (bases 1 to 7041)
  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: "PLoS One.";
            date: "2016-05-2"; volume: "11(5):e0154684. doi"; pages: " 
            10.1371/journal.pone.0154684. eCollection 2016"
COMMENT     Sequence Label: pMA3790
FEATURES             Location/Qualifiers
     source          1..7041
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        115..418
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        419..622
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     573..593
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             762..791
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     polyA_signal    1600..1721
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1637..1656)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1691..1710
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     polyA_signal    1779..2003
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     1992..2009
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     promoter        2029..2047
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     2029..2046
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     CDS             complement(join(2153..2866,2867..2869,2870..2872))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(2156..2866,2867..2869,2870..2872))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2198..2219)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2547..2566
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2806..2827
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      2869..2878
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(2894..4072)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          2903..3841
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     complement(2926..2946)
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     rep_origin      complement(4083..4538)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4220..4241)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4432..4451
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        4565..4669
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4694..4714)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        4738..4948
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        4760..4956
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4807..4942
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4991..5785
                     /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             4991..5785
                     /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(5045..5064)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5655..5674
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5973..5992)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6017..6064
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      6393..6981
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6882..6901
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgccat gcattagtta
       61 ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac
      121 ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc
      181 aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt
      241 ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac
      301 gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac
      361 cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt
      421 gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac ggggatttcc
      481 aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt
      541 tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg
      601 ggaggtctat ataagcagag ctggtttagt gaaccgtcag atccgctaga tcgaattccg
      661 ccaccatgct gtttaatctg aggattctgt taaacaatgc agcttttaga aatggtcaca
      721 acttcatggt tcgaaatttt cggtgtggac aaccactaca agagcagaaa ctcatcagcg
      781 aggaggacct gggatctgcc cgcaacgtgc ccgtgggctt tggagagagc tggaagaagc
      841 acctcagcgg ggagttcggg aaaccgtatt ttatcaagct aatgggattt gttgcagaag
      901 aaagaaagca ttacactgtt tatccacccc cacaccaagt cttcacctgg acccagatgt
      961 gtgacataaa agatgtgaag gttgtcatcc tgggacagga tccagcccat ggacctaatc
     1021 aagctcacgg gctctgcttt agtgttcaaa ggcctgttcc gcctccgccc agtttggaga
     1081 acatttataa agagttgtct acagacatag aggattttgt tcatcctggc catggagatt
     1141 tatctgggtg ggccaagcaa ggtgttctcc ttctcaacgc tgtcctcacg gttcgtgccc
     1201 atcaagccaa ctctcataag gagcgaggct gggagcagtt cactgatgca gttgtgtcct
     1261 ggctaaatca gaactcgaat ggccttgttt tcttgctctg gggctcttat gctcagaaga
     1321 agggcagtgc cattgatagg aagcggcacc atgtactaca gacggctcat ccctcccctt
     1381 tgtcagtgta tagagggttc tttggatgta gacacttttc aaagaccaat gagctgctgc
     1441 agaagtctgg caagaagccc attgactgga aggagctgtg agtctagata actgatcata
     1501 atcagccata ccacatttgt agaggtttta cttgctttaa aaaacctccc acacctcccc
     1561 ctgaacctga aacataaaat gaatgcaatt gttgttgtta acttgtttat tgcagcttat
     1621 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     1681 cattctagtt gtggtttgtc caaactcatc aatgtatctt aacgcgcgtc aattcgaggg
     1741 gctgcaggaa ttcgatctgg ttctttccgc ctcagaagcc atagagccca ccgcatcccc
     1801 agcatgcctg ctattgtctt cccaatcctc ccccttgctg tcctgcccca ccccaccccc
     1861 cagaatagaa tgacacctac tcagacaatg cgatgcaatt tcctcatttt attaggaaag
     1921 gacagtggga gtggcacctt ccagggtcaa ggaaggcacg ggggaggggc aaacaacaga
     1981 tggctggcaa ctagaaggca cagtcgaggc tgatcagcga gctctagcat ttaggtgaca
     2041 ctatagaata gggccctcta gaactagtag gcctattatt tttgacacca gacaagttgg
     2101 taatggtagc gaccggcgct cagttggaat tctagagtcg cggccggccg ctttacttgt
     2161 acagctcgtc catgccgaga gtgatcccgg cggcggtcac gaactccagc aggaccatgt
     2221 gatcgcgctt ctcgttgggg tctttgctca gggcggactg ggtgctcagg tagtggttgt
     2281 cgggcagcag cacggggccg tcgccgatgg gggtgttctg ctggtagtgg tcggcgagct
     2341 gcacgctgcc gtcctcgatg ttgtggcgga tcttgaagtt caccttgatg ccgttcttct
     2401 gcttgtcggc catgatatag acgttgtggc tgttgtagtt gtactccagc ttgtgcccca
     2461 ggatgttgcc gtcctccttg aagtcgatgc ccttcagctc gatgcggttc accagggtgt
     2521 cgccctcgaa cttcacctcg gcgcgggtct tgtagttgcc gtcgtccttg aagaagatgg
     2581 tgcgctcctg gacgtagcct tcgggcatgg cggacttgaa gaagtcgtgc tgcttcatgt
     2641 ggtcggggta gcggctgaag cactgcacgc cgtaggtcag ggtggtcacg agggtgggcc
     2701 agggcacggg cagcttgccg gtggtgcaga tgaacttcag ggtcagcttg ccgtaggtgg
     2761 catcgccctc gccctcgccg gacacgctga acttgtggcc gtttacgtcg ccgtccagct
     2821 cgaccaggat gggcaccacc ccggtgaaca gctcctcgcc cttgctcacc atggtggcga
     2881 ccggtaccaa gcttcacgac acctgaaatg gaagaaaaaa actttgaacc actgtctgag
     2941 gcttgagaat gaaccaagat ccaaactcaa aaagggcaaa ttccaaggag aattacatca
     3001 agtgccaagc tggcctaact tcagtctcca cccactcagt gtggggaaac tccatcgcat
     3061 aaaacccctc cccccaacct aaagacgacg tactccaaaa gctcgagaac taatcgaggt
     3121 gcctggacgg cgcccggtac tccgtggagt cacatgaagc gacggctgag gacggaaagg
     3181 cccttttcct ttgtgtgggt gactcacccg cccgctctcc cgagcgccgc gtcctccatt
     3241 ttgagctccc tgcagcaggg ccgggaagcg gccatctttc cgctcacgca actggtgccg
     3301 accgggccag ccttgccgcc cagggcgggg cgatacacgg cggcgcgagg ccaggcacca
     3361 gagcaggccg gccagcttga gactaccccc gtccgattct cggtggccgc gctcgcaggc
     3421 cccgcctcgc cgaacatgtg cgctgggacg cacgggcccc gtcgccgccc gcggccccaa
     3481 aaaccgaaat accagtgtgc agatcttggc ccgcatttac aagactatct tgccagaaaa
     3541 aaagcgtcgc agcaggtcat caaaaatttt aaatggctag agacttatcg aaagcagcga
     3601 gacaggcgcg aaggtgccac cagattcgca cgcggcggcc ccagcgccca ggccaggcct
     3661 caactcaagc acgaggcgaa ggggctcctt aagcgcaagg cctcgaactc tcccacccac
     3721 ttccaacccg aagctcggga tcaagaatca cgtactgcag ccaggtggaa gtaattcaag
     3781 gcacgcaagg gccataaccc gtaaagaggc caggcccgcg ggaaccacac acggcactta
     3841 cctgtgttct ggcggcaaac ccgttgcgaa aaagaacgtt cacggcgact actgcactta
     3901 tatacggttc tcccccaccc tcgggaaaaa ggcggagcca gtacacgaca tcactttccc
     3961 agtttacccc gcgccacctt ctctaggcac cggttcaatt gccgacccct ccccccaact
     4021 tctcggggac tgtgggcgat gtgcgctctg cccactgacg ggcaccggag cctcacgcgc
     4081 gtaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
     4141 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
     4201 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
     4261 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
     4321 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
     4381 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
     4441 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
     4501 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtca ggtggcactt ttcggggaaa
     4561 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     4621 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagtc ctgaggcgga
     4681 aagaaccagc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     4741 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact
     4801 ccgcccatcc cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta
     4861 atttttttta tttatgcaga ggccgaggcc gcctcggcct ctgagctatt ccagaagtag
     4921 tgaggaggct tttttggagg cctaggcttt tgcaaagatc gatcaagaga caggatgagg
     4981 atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga
     5041 gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt
     5101 ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct
     5161 gaatgaactg caagacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg
     5221 cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt
     5281 gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc
     5341 tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc
     5401 gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga
     5461 tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgag
     5521 catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat
     5581 ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg
     5641 ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc
     5701 tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta
     5761 tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg
     5821 acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     5881 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     5941 gagttcttcg cccaccctag ggggaggcta actgaaacac ggaaggagac aataccggaa
     6001 ggaacccgcg ctatgacggc aataaaaaga cagaataaaa cgcacggtgt tgggtcgttt
     6061 gttcataaac gcggggttcg gtcccagggc tggcactctg tcgatacccc accgagaccc
     6121 cattggggcc aatacgcccg cgtttcttcc ttttccccac cccacccccc aagttcgggt
     6181 gaaggcccag ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc ctcaggttac
     6241 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
     6301 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     6361 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     6421 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     6481 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
     6541 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     6601 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     6661 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     6721 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     6781 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     6841 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     6901 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     6961 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     7021 tgctggcctt ttgctcacat g
//
