LOCUS       sequence-103422-        6625 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  mtZFN2G architecture bearing (+) FokI domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6625)
  AUTHORS   Gammage PA, Rorbach J, Vincent AI, Rebar EJ, Minczuk M
  TITLE     Mitochondrially targeted ZFNs for selective degradation of 
            pathogenic mitochondrial genomes bearing large-scale deletions or 
            point mutations.
  JOURNAL   EMBO Mol Med. 2014 Apr;6(4):458-66. doi: 10.1002/emmm.201303672. 
            Epub 2014 Feb 24.
  PUBMED    24567072
REFERENCE   2  (bases 1 to 6625)
  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: 
            "10.1002/emmm.201303672"; journalName: "EMBO Mol Med"; date: 
            "2014-04"; volume: "6"; issue: "4"; pages: "458-66"
COMMENT     Sequence Label: pcDNA3.1(-)_mtZFN(+)2G
FEATURES             Location/Qualifiers
     source          1..6625
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1068..1094
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YPYDVPDYA"
     CDS             1068..1094
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(2219..2236)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2225..2449
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2495..2923
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2582..2601)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2792..2813
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2932..2952)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2937..3266
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3117..3252
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3179..3198
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3333..4127
                     /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
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3387..3406)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3997..4016
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4301..4422
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4338..4357)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4392..4411
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4471..4487)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4471..4487)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4484..4506)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4495..4511
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4519..4525,4526..4543,4544..4549))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4564..4585
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4702..4719)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4873..5458)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4953..4972)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5629..6420,6421..6489))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6252..6271
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6490..6594)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaacgg gccctctaga atgttggggt ttgtgggtcg ggtggccgct gctccggcct
      961 ccggggcctt gcggagactc accccttcag cgtcgctgcc cccagctcag ctcttactgc
     1021 gggccgctcc gacggcggtc catcctgtca gggactatgc ggcgcaatac ccctacgacg
     1081 tgcccgacta cgccgtggat gaaatgacca aaaagttcgg cacgctcacc attcacgaca
     1141 ccgaaaaggc cgccgaattc atggctgaga ggcccttcca gtgtcgaatc tgcatgcgta
     1201 acttcagtga ccgctccgcc ctgtcccgcc acatccgcac ccacaccggc gagaagcctt
     1261 ttgcctgtga catttgtggg aggaaatttg cccgctccga cgccctggcc cgccatacca
     1321 agatacacac gggcagccaa aagcccttcc agtgtcgaat ctgcatgcgt aacttcagtc
     1381 gctccgacac cctgtccgag cacatccgca cccacaccgg cgagaagcct tttgcctgtg
     1441 acatttgtgg gaggaaattt gcccaccact ccgcccgccg ctgtcatacc aagatacaca
     1501 cgggcgagaa gcccttccag tgtcgaatct gcatgcgtaa gtttgcccgc tccgacaacc
     1561 tgacccgcca taccaagata cacctgcggg gatcccagct ggtgaagagc gagctggagg
     1621 agaagaagtc cgagctgcgg cacaagctga agtacgtgcc ccacgagtac atcgagctga
     1681 tcgagatcgc caggaacagc acccaggacc gcatcctgga gatgaaggtg atggagttct
     1741 tcatgaaggt gtacggctac aggggaaagc acctgggcgg aagcagaaag cctgacggcg
     1801 ccatctatac agtgggcagc cccatcgatt acggcgtgat cgtggacaca aaggcctaca
     1861 gcggcggcta caatctgcct atcggccagg ccgacgagat gcagagatac gtgaaggaga
     1921 accagacccg gaataagcac atcaacccca acgagtggtg gaaggtgtac cctagcagcg
     1981 tgaccgagtt caagttcctg ttcgtgagcg gccacttcaa gggcaactac aaggcccagc
     2041 tgaccaggct gaaccgcaag accaactgca atggcgccgt gctgagcgtg gaggagctgc
     2101 tgatcggcgg cgagatgatc aaagccggca ccctgacact ggaggaggtg cggcgcaagt
     2161 tcaacaacgg cgagatcaac ttctgaggta ccaagcttaa gtttaaaccg ctgatcagcc
     2221 tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg
     2281 accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat
     2341 tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag
     2401 gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg
     2461 gaaagaacca gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc
     2521 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
     2581 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     2641 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     2701 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     2761 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     2821 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
     2881 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attaattctg tggaatgtgt
     2941 gtcagttagg gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc
     3001 atctcaatta gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta
     3061 tgcaaagcat gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
     3121 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     3181 tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag tgaggaggct
     3241 tttttggagg cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc attttcggat
     3301 ctgatcaaga gacaggatga ggatcgtttc gcatgattga acaagatgga ttgcacgcag
     3361 gttctccggc cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg
     3421 gctgctctga tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca
     3481 agaccgacct gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc
     3541 tggccacgac gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg
     3601 actggctgct attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg
     3661 ccgagaaagt atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta
     3721 cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag
     3781 ccggtcttgt cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac
     3841 tgttcgccag gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg acccatggcg
     3901 atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg
     3961 gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg
     4021 aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg
     4081 attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg
     4141 gttcgaaatg accgaccaag cgacgcccaa cctgccatca cgagatttcg attccaccgc
     4201 cgccttctat gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct
     4261 ccagcgcggg gatctcatgc tggagttctt cgcccacccc aacttgttta ttgcagctta
     4321 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     4381 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc
     4441 gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta
     4501 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     4561 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     4621 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     4681 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     4741 gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat caggggataa
     4801 cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     4861 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     4921 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     4981 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     5041 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     5101 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     5161 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     5221 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     5281 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     5341 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     5401 tggtagcggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     5461 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     5521 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     5581 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     5641 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     5701 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     5761 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     5821 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     5881 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     5941 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     6001 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     6061 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     6121 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     6181 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     6241 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     6301 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     6361 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     6421 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     6481 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     6541 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     6601 tccccgaaaa gtgccacctg acgtc
//
