LOCUS       sequence-435508-        5262 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of apt-CBE-3'end protein 
            (MCP_32aa-linker_evoAPOBEC1) with P2A-EGFP marker .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5262)
  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 5262)
  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-CBE-3'end
FEATURES             Location/Qualifiers
     source          1..5262
                     /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             1927..2175
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             2206..2454
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             2497..2517
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2526..2543
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2566..2583)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2572..2796
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2867..2883)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2867..2883)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2880..2902)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2891..2907
                     /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(2915..2921,2922..2939,2940..2945))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2960..2981
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3098..3115)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3269..3857)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3349..3368)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4028..4819,4820..4888))
                     /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     4651..4670
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4889..4993)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5072..5091)
                     /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 atcctctggc
     1081 agcgagacac caggaacaag cgagtcagca acaccagaga gcagtggcgg cagcagcggc
     1141 ggcagcagtt caaagactgg gcctgtcgcc gtcgatccaa ccctgcgccg ccggattgaa
     1201 cctcacgagt ttgaagtgtt ctttgacccc cgggagctga gaaaggagac atgcctgctg
     1261 tacgagatca actggggagg caggcactcc atctggaggc acacctctca gaacacaaat
     1321 aagcacgtgg aggtgaactt catcgagaag tttaccacag agcggtactt ctgccccaat
     1381 accagatgta gcatcacatg gtttctgagc tggtcccctt gcggagagtg tagcagggcc
     1441 atcaccgagt tcctgtccag atatccaaat gtgacactgt ttatctacat cgccaggctg
     1501 tatcacctgg caaacccaag gaataggcag ggcctgcgcg atctgatcag ctccggcgtg
     1561 accatccaga tcatgacaga gcaggagtcc ggctactgct ggcacaactt cgtgaattat
     1621 tctcctagca acgagtccca ctggcctagg tacccacacc tgtgggtgcg cctgtacgtg
     1681 ctggagctgt attgcatcat cctgggcctg cccccttgtc tgaatatcct gcggagaaag
     1741 cagagccagc tgacctcctt tacaatcgcc ctgcagtctt gtcactatca gaggctgcca
     1801 ccccacatcc tgtgggccac aggcctgaag tctggcggat ctagcggagg atcctctggc
     1861 agcgagacac caggaacaag cgagtcagca acaccagaga gcagtggcgg cagcagcggc
     1921 ggcagcacta atctgagcga catcattgag aaggagactg ggaaacagct ggtcattcag
     1981 gagtccatcc tgatgctgcc tgaggaggtg gaggaagtga tcggcaacaa gccagagtct
     2041 gacatcctgg tgcacaccgc ctacgacgag tccacagatg agaatgtgat gctgctgacc
     2101 tctgacgccc ccgagtataa gccttgggcc ctggtcatcc aggattctaa cggcgagaat
     2161 aagatcaaga tgctgagcgg aggatccgga ggatctggag gcagcaccaa cctgtctgac
     2221 atcatcgaga aggagacagg caagcagctg gtcatccagg agagcatcct gatgctgccc
     2281 gaagaagtcg aagaagtgat cggaaacaag cctgagagcg atatcctggt ccataccgcc
     2341 tacgacgaga gtaccgacga aaatgtgatg ctgctgacat ccgacgcccc agagtataag
     2401 ccctgggctc tggtcatcca ggattccaac ggagagaaca aaatcaaaat gctgtctggc
     2461 ggctcaaaaa gaaccgccga cggcagcgaa ttcgagccca agaagaagag gaaagtctaa
     2521 ccggtcatca tcaccatcac cattgagttt aaacccgctg atcagcctcg actgtgcctt
     2581 ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg
     2641 ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt
     2701 gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca
     2761 atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct
     2821 ggggctcgat accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc
     2881 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     2941 gtaaagccta ggatgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     3001 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     3061 gaagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     3121 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     3181 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     3241 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     3301 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     3361 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     3421 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     3481 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     3541 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     3601 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     3661 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     3721 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     3781 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     3841 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac actcagtgga
     3901 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     3961 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     4021 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     4081 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     4141 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     4201 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     4261 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     4321 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     4381 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     4441 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     4501 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     4561 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     4621 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     4681 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     4741 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     4801 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     4861 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     4921 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     4981 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc
     5041 gatctcccga tcccctaggg tcgactctca gtacaatctg ctctgatgcc gcatagttaa
     5101 gccagtatct gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg agcaaaattt
     5161 aagctacaac aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt agggttaggc
     5221 gttttgcgct gcttcgcgat gtacgggcca gatatacgcg tt
//
