LOCUS       sequence-450044-        4296 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of apt-ABE8.20 protein 
            (TadA8.20_32aa-linker_Com) .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4296)
  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 4296)
  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: apt-ABE8.20
FEATURES             Location/Qualifiers
     source          1..4296
                     /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"
     CDS             694..714
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             718..1215
                     /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             1531..1551
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1560..1577
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(1600..1617)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1606..1830
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(1901..1917)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1901..1917)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1914..1936)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1925..1941
                     /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(1949..1955,1956..1973,1974..1979))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1994..2015
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2132..2149)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2303..2891)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2383..2402)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3062..3853,3854..3922))
                     /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     3685..3704
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3923..4027)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4106..4125)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatattga 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 aaacggacag ccgacggaag cgagttcgag tcaccaaaga agaagcggaa agtcagctct
      721 gaggtggagt tttcccatga gtactggatg agacacgcat tgactctcgc aaagagggct
      781 cgagatgaac gcgaggtgcc cgtgggggca gtactcgtgc tcaacaatcg cgtaatcggc
      841 gaaggttgga atagggcaat cggactccac gaccccactg cacatgcgga aatcatggcc
      901 cttcgacagg gagggcttgt gatgcagaat tatcgacttt atgatgcgac gctgtactcg
      961 acgtttgaac cttgcgtaat gtgcgcggga gctatgattc actcccgcat tggacgagtt
     1021 gtattcggtg ttcgcaacgc caagacgggt gccgcaggtt cactgatgga cgtgctgcat
     1081 catccaggca tgaaccaccg ggtagaaatc acagaaggca tattggcgga cgaatgtgcg
     1141 gcgctgttgt gtcgtttttt tcgcatgccc aggcgggtct ttaacgccca gaaaaaagca
     1201 caatcctcta ctgactctgg tggttcttct ggtggttcta gcggcagcga gactcccggg
     1261 acctcagagt ccgccacacc cgaaagttct ggtggttctt ctggtggttc taaaagcatc
     1321 agatgcaaga actgcaacaa gctgctgttc aaggccgaca gcttcgacca catcgagatc
     1381 cggtgtccta gatgtaaaag acacatcatc atgctgaacg cctgcgagca ccccaccgag
     1441 aagcactgcg gcaagcggga aaagatcacc cacagcgacg agacagtgcg gtacggttca
     1501 aaaagaaccg ccgacggcag cgaattcgag cccaagaaga agaggaaagt ctaaccggtc
     1561 atcatcacca tcaccattga gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt
     1621 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     1681 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
     1741 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca
     1801 ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct
     1861 cgataccgtc gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt
     1921 gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag
     1981 cctagggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt
     2041 tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag
     2101 gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     2161 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     2221 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     2281 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     2341 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     2401 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     2461 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     2521 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     2581 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     2641 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     2701 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     2761 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     2821 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     2881 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacactcag tggaacgaaa
     2941 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     3001 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     3061 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     3121 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     3181 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     3241 accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc
     3301 agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca
     3361 acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat
     3421 tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag
     3481 cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac
     3541 tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt
     3601 ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt
     3661 gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact ttaaaagtgc
     3721 tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg ctgttgagat
     3781 ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt actttcacca
     3841 gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga ataagggcga
     3901 cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc atttatcagg
     3961 gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa caaatagggg
     4021 ttccgcgcac atttccccga aaagtgccac ctgacgtcga cggatcggga gatcgatctc
     4081 ccgatcccct agggtcgact ctcagtacaa tctgctctga tgccgcatag ttaagccagt
     4141 atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta
     4201 caacaaggca aggcttgacc gacaattgca tgaagaatct gcttagggtt aggcgttttg
     4261 cgctgcttcg cgatgtacgg gccagatata cgcgtt
//
