LOCUS       sequence-375371-        6041 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expression of mitochondrial v8 ZF-DdCBE in mammalian cells 
            (N-terminal DddAC, Left).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6041)
  AUTHORS   Willis JCW, Silva-Pinheiro P, Widdup L, Minczuk M, Liu DR
  TITLE     Compact zinc finger base editors that edit mitochondrial or nuclear 
            DNA in vitro and in vivo.
  JOURNAL   Nat Commun. 2022 Nov 23;13(1):7204. doi: 10.1038/s41467-022-34784-7.
  PUBMED    36418298
REFERENCE   2  (bases 1 to 6041)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2022-11-23"; volume: "13(1):7204. doi"; pages: " 
            10.1038/s41467-022-34784-7"
COMMENT     Sequence Label: pB7N_LT51-Mt-tk_v8_AGKS
FEATURES             Location/Qualifiers
     source          1..6041
                     /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             1051..1074
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             1771..2022
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     rep_origin      2273..2701
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2360..2379)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2570..2591
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2710..2730)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2715..3044
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2895..3030
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2957..2976
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3092..3139
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3158..3556
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    3714..3835
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3751..3770)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3805..3824
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3884..3900)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3884..3900)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3897..3919)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3908..3924
                     /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(3932..3938,3939..3956,3957..3962))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3977..3998
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4115..4132)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4286..4874)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4366..4385)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5045..5836,5837..5905))
                     /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     5668..5687
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5906..6010)
                     /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 tatgacctta 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 accatgttgg ggtttgtggg tcgggtggcc gctgctccgg cctccggggc cttgcggaga
      961 ctcacccctt cagcgtcgct gcccccagct cagctcttac tgcgggccgc tccgacggcg
     1021 gtccatcctg tcagggacta tgcggcgcaa gattacaagg acgacgatga caaggtggat
     1081 gaaatgacca aaaagttcgg cacgctcacc attcacgaca ccgaaaaggg atcacttcag
     1141 aaaaagctgg aggaactgga gttggacgcc gcttctggcg gctcagccat tccagtgaag
     1201 cgcggcgcta ccggtgaaac caaagtgttt atcggtaaca gcaacagccc gaagagcccg
     1261 accaaaggcg gttgcggatc cggcggagga ggttctggcg gttctggagg aagtatggct
     1321 gagaggccct atgcttgccc tgagtgcggt aaaagctttt ctcaactcgc tcatctgcga
     1381 gcacacatcc gcacccacac cggcgagaag ccttacgcat gtccagagtg tgggaagtca
     1441 ttctctacaa gcggctctct cgttagacat attcgcaccc atacaggaga aaagccctat
     1501 gcctgccctg agtgcggaaa gtcctttagc cagagagcta atctcagggc ccacataagg
     1561 acacatactg gggagaaacc ttacgcgtgt ccggaatgtg gcaagagttt ttccacttct
     1621 ggaagcttgg tgaggcatat cagaacccac actggcgaaa agccgtacgc ttgcccagaa
     1681 tgcggtaagt ccttctctca agcctctaac ttgattagcc ataccaagat acacctgcgg
     1741 agcggaggat ccggaggatc tggaggcagc actaatctga gcgacatcat tgagaaggag
     1801 actgggaaac agctggtcat tcaggagtcc atcctgatgc tgcctgagga ggtggaggaa
     1861 gtgatcggca acaagccaga gtctgacatc ctggtgcaca ccgcctacga cgagtccaca
     1921 gatgagaatg tgatgctgct gacctctgac gcccccgagt ataagccttg ggccctggtc
     1981 atccaggatt ctaacggcga gaataagatc aagatgctgt gataaaccac gatcgttatg
     2041 ctgatcatac cctaatgatc ccagcaagat aatgtcctgt cttctaagat gtgcatcaag
     2101 cctggtacat actgaaaacc ctataaggtc ctggataatt tttgtttgat tattcattga
     2161 agaaacattt attttccaat tgtgtgaagt ttttgactgt taataaaaga atctgtcaac
     2221 catcaaactt ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc
     2281 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     2341 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     2401 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     2461 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
     2521 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     2581 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
     2641 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     2701 taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca
     2761 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct
     2821 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc
     2881 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg
     2941 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc
     3001 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt
     3061 gtatatccat tttcggatct gatcagcacg tgttgacaat taatcatcgg catagtatat
     3121 cggcatagta taatacgaca aggtgaggaa ctaaaccatg gccaagcctt tgtctcaaga
     3181 agaatccacc ctcattgaaa gagcaacggc tacaatcaac agcatcccca tctctgaaga
     3241 ctacagcgtc gccagcgcag ctctctctag cgacggccgc atcttcactg gtgtcaatgt
     3301 atatcatttt actgggggac cttgtgcaga actcgtggtg ctgggcactg ctgctgctgc
     3361 ggcagctggc aacctgactt gtatcgtcgc gatcggaaat gagaacaggg gcatcttgag
     3421 cccctgcgga cggtgccgac aggtgcttct cgatctgcat cctgggatca aagccatagt
     3481 gaaggacagt gatggacagc cgacggcagt tgggattcgt gaattgctgc cctctggtta
     3541 tgtgtgggag ggctaagcac ttcgtggccg aggagcagga ctgacacgtg ctacgagatt
     3601 tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg
     3661 gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac cccaacttgt
     3721 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     3781 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg
     3841 tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg
     3901 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     3961 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     4021 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     4081 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     4141 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     4201 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     4261 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     4321 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     4381 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     4441 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     4501 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     4561 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     4621 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     4681 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     4741 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     4801 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     4861 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     4921 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     4981 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     5041 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     5101 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     5161 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     5221 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     5281 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     5341 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     5401 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     5461 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     5521 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     5581 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     5641 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     5701 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     5761 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     5821 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     5881 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     5941 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     6001 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt c
//
