LOCUS       sequence-440937-        6171 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of MOBE4 protein 
            (apt-CBE-SL3_PT2A_apt-ABE8.20).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6171)
  AUTHORS   Cowan QT, Gu S, Gu W, Ranzau BL, Simonson TS, Komor AC
  TITLE     Development of multiplexed orthogonal base editor (MOBE) systems.
  JOURNAL   Nat Biotechnol. 2024 May 21. doi: 10.1038/s41587-024-02240-0.
  PUBMED    38773305
REFERENCE   2  (bases 1 to 6171)
  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 
            Biotechnol. 2024 May 21. doi: 10.1038/s41587-024-02240-0."
COMMENT     Sequence Label: MOBE4
FEATURES             Location/Qualifiers
     source          1..6171
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      652..661
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             661..1047
                     /codon_start=1
                     /product="bacteriophage MS2 coat protein"
                     /label=MS2-N55K
                     /note="binds with high affinity to a specific stem-loop 
                     structure in the viral RNA (Lim et al., 1994)"
                     /translation="ASNFTQFVLVDNGGTGDVTVAPSNFANGVAEWISSNSRSQAYKVT
                     CSVRQSSAQKRKYTIKVEVPKVATQTVGGVELPVAAWRSYLNMELTIPIFATNSDCELI
                     VKAMQGLLKDGNPIPSAIAANSGIY"
     CDS             1813..2061
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             2092..2340
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             2383..2403
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2413..2469
                     /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"
     CDS             2479..2532
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             2569..2589
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2593..3090
                     /codon_start=1
                     /product="DNA adenosine deaminase evolved from the E. coli 
                     tRNA adenosine deaminase TadA (Gaudelli et al., 2017)"
                     /label=TadA*(7.10)
                     /translation="SEVEFSHEYWMRHALTLAKRARDEREVPVGAVLVLNNRVIGEGWN
                     RAIGLHDPTAHAEIMALRQGGLVMQNYRLYDATLYSTFEPCVMCAGAMIHSRIGRVVFG
                     VRNAKTGAAGSLMDVLHHPGMNHRVEITEGILADECAALLCRFFRMPRRVFNAQKKAQS
                     STD"
     CDS             3406..3426
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3435..3452
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(3475..3492)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3481..3705
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3776..3792)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3776..3792)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3789..3811)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3800..3816
                     /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(3824..3830,3831..3848,3849..3854))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3869..3890
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4007..4024)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4178..4766)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4258..4277)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4937..5728,5729..5797))
                     /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     5560..5579
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5798..5902)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5981..6000)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc
      601 agatccgcta gagatccgcg gccgctaata cgactcacta tagggagagc cgccaccatg
      661 gcaagtaact tcactcaatt tgtgcttgta gacaacggag gtaccggaga tgtaacagta
      721 gcacccagta atttcgcaaa cggcgtggcg gaatggatta gttcaaattc aagatctcag
      781 gcctataagg tcacttgctc cgtcaggcag tctagtgccc aaaagcgcaa atacacgatt
      841 aaagtagagg taccgaaggt cgctacgcaa actgttgggg gagttgaact tccagtggct
      901 gcttggcgct catacctgaa catggaactg actatcccca tattcgctac taactccgat
      961 tgtgaactca tagtaaaagc tatgcaggga ttgctcaagg atggcaaccc cattccaagt
     1021 gccatagcag cgaactccgg gatatattct tctggaggat ctagcggagg atccagtggc
     1081 ggcagcagcg gcggcagcag ttcaaagact gggcctgtcg ccgtcgatcc aaccctgcgc
     1141 cgccggattg aacctcacga gtttgaagtg ttctttgacc cccgggagct gagaaaggag
     1201 acatgcctgc tgtacgagat caactgggga ggcaggcact ccatctggag gcacacctct
     1261 cagaacacaa ataagcacgt ggaggtgaac ttcatcgaga agtttaccac agagcggtac
     1321 ttctgcccca ataccagatg tagcatcaca tggtttctga gctggtcccc ttgcggagag
     1381 tgtagcaggg ccatcaccga gttcctgtcc agatatccaa atgtgacact gtttatctac
     1441 atcgccaggc tgtatcacct ggcaaaccca aggaataggc agggcctgcg cgatctgatc
     1501 agctccggcg tgaccatcca gatcatgaca gagcaggagt ccggctactg ctggcacaac
     1561 ttcgtgaatt attctcctag caacgagtcc cactggccta ggtacccaca cctgtgggtg
     1621 cgcctgtacg tgctggagct gtattgcatc atcctgggcc tgcccccttg tctgaatatc
     1681 ctgcggagaa agcagagcca gctgacctcc tttacaatcg ccctgcagtc ttgtcactat
     1741 cagaggctgc caccccacat cctgtgggcc acaggcctga agtctggcgg atctggaggc
     1801 tccggcggca gcactaatct gagcgacatc attgagaagg agactgggaa acagctggtc
     1861 attcaggagt ccatcctgat gctgcctgag gaggtggagg aagtgatcgg caacaagcca
     1921 gagtctgaca tcctggtgca caccgcctac gacgagtcca cagatgagaa tgtgatgctg
     1981 ctgacctctg acgcccccga gtataagcct tgggccctgg tcatccagga ttctaacggc
     2041 gagaataaga tcaagatgct gagcggagga tccggaggat ctggaggcag caccaacctg
     2101 tctgacatca tcgagaagga gacaggcaag cagctggtca tccaggagag catcctgatg
     2161 ctgcccgaag aagtcgaaga agtgatcgga aacaagcctg agagcgatat cctggtccat
     2221 accgcctacg acgagagtac cgacgaaaat gtgatgctgc tgacatccga cgccccagag
     2281 tataagccct gggctctggt catccaggat tccaacggag agaacaaaat caaaatgctg
     2341 tctggcggct caaaaagaac cgccgacggc agcgaattcg agcccaagaa gaagaggaaa
     2401 gtcggaagcg gagctactaa cttcagcctg ctgaagcagg ctggagacgt ggaggagaac
     2461 cctggacctg gcagtggtga gggcagagga agtctgctaa catgcggtga cgtcgaggag
     2521 aatcctggcc caatgaaacg gacagccgac ggaagcgagt tcgagtcacc aaagaagaag
     2581 cggaaagtca gctctgaggt ggagttttcc catgagtact ggatgagaca cgcattgact
     2641 ctcgcaaaga gggctcgaga tgaacgcgag gtgcccgtgg gggcagtact cgtgctcaac
     2701 aatcgcgtaa tcggcgaagg ttggaatagg gcaatcggac tccacgaccc cactgcacat
     2761 gcggaaatca tggcccttcg acagggaggg cttgtgatgc agaattatcg actttatgat
     2821 gcgacgctgt actcgacgtt tgaaccttgc gtaatgtgcg cgggagctat gattcactcc
     2881 cgcattggac gagttgtatt cggtgttcgc aacgccaaga cgggtgccgc aggttcactg
     2941 atggacgtgc tgcatcatcc aggcatgaac caccgggtag aaatcacaga aggcatattg
     3001 gcggacgaat gtgcggcgct gttgtgtcgt ttttttcgca tgcccaggcg ggtctttaac
     3061 gcccagaaaa aagcacaatc ctctactgac tctggtggtt cttctggtgg ttctagcggc
     3121 agcgagactc ccgggacctc agagtccgcc acacccgaaa gttctggtgg ttcttctggt
     3181 ggttctaaaa gcatcagatg caagaactgc aacaagctgc tgttcaaggc cgacagcttc
     3241 gaccacatcg agatccggtg tcctagatgt aaaagacaca tcatcatgct gaacgcctgc
     3301 gagcacccca ccgagaagca ctgcggcaag cgggaaaaga tcacccacag cgacgagaca
     3361 gtgcggtacg gttcaaaaag aactgccgac ggctctgagt ttgaacccaa gaagaaaagg
     3421 aaagtctaac cggtcatcat caccatcacc attgagttta aacccgctga tcagcctcga
     3481 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     3541 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     3601 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     3661 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa
     3721 gaaccagctg gggctcgata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     3781 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     3841 gcataaagtg taaagcctag gatgcctaat gagtgagcta actcacatta attgcgttgc
     3901 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     3961 aacgcgcggg aagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     4021 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     4081 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     4141 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     4201 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     4261 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     4321 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     4381 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     4441 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     4501 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     4561 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     4621 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     4681 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     4741 ttacgcgcag aaaaaaagga tctcaagaaa atcctttgat cttttctacg gggtctgaca
     4801 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     4861 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     4921 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     4981 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     5041 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     5101 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     5161 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     5221 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     5281 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     5341 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     5401 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     5461 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     5521 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     5581 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     5641 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     5701 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     5761 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     5821 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     5881 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat
     5941 cgggagatcg atctcccgat cccctagggt cgactctcag tacaatctgc tctgatgccg
     6001 catagttaag ccagtatctg ctccctgctt gtgtgttgga ggtcgctgag tagtgcgcga
     6061 gcaaaattta agctacaaca aggcaaggct tgaccgacaa ttgcatgaag aatctgctta
     6121 gggttaggcg ttttgcgctg cttcgcgatg tacgggccag atatacgcgt t
//
