LOCUS       sequence-270437-        5466 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the C-terminal split-intein fragment of S. aureus BE3.9max
            (KKH variant) from the CMV promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5466)
  AUTHORS   Levy JM, Yeh WH, Pendse N, Davis JR, Hennessey E, Butcher R, Koblan 
            LW, Comander J, Liu Q, Liu DR
  TITLE     Cytosine and adenine base editing of the brain, liver, retina, heart
            and skeletal muscle of mice via adeno-associated viruses.
  JOURNAL   Nat Biomed Eng. 2020 Jan;4(1):97-110. doi: 
            10.1038/s41551-019-0501-5. Epub 2020 Jan 14.
  PUBMED    31937940
REFERENCE   2  (bases 1 to 5466)
  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: 
            "10.1038/s41551-019-0501-5"; journalName: "Nat Biomed Eng"; date: 
            "2020-01"; volume: "4"; issue: "1"; pages: "97-110"
COMMENT     Sequence Label: CMV_Npu-saBE3.9max_KKH C-terminal
FEATURES             Location/Qualifiers
     source          1..5466
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        132..335
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     286..306
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     377..396
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        377..395
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             445..465
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2161..2409
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             2452..2472
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2481..2498
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2521..2538)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2527..2751
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2822..2838)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2822..2838)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2835..2857)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2846..2862
                     /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(2870..2876,2877..2894,2895..2900))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2915..2936
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3053..3070)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3224..3812)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3304..3323)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3983..4774,4775..4843))
                     /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     4606..4625
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4844..4948)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5027..5046)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        join(5218..5466,1..131)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg
       61 cccagtacat gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg
      121 ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact
      181 cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa
      241 atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta
      301 ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt cagatccgct
      361 agagatccgc ggccgctaat acgactcact atagggagag ccgccaccat gaaacggaca
      421 gccgacggaa gcgagttcga gtcaccaaag aagaagcgga aagtcatcaa gattgctaca
      481 cggaaatacc tgggaaagca gaacgtgtac gacatcggcg tggagcggga tcacaacttc
      541 gccctgaaga atggctttat cgccagcaat tgtctgtatt ctctggaggc catccctctg
      601 gaggacctgc tgaacaatcc attcaactac gaggtggatc acatcatccc ccggagcgtg
      661 agcttcgaca attcttttaa caataaggtg ctggtgaagc aggaggagaa cagcaagaag
      721 ggcaatagga cccctttcca gtacctgtct agctccgatt ctaagatcag ctacgagaca
      781 ttcaagaagc acatcctgaa tctggccaag ggcaagggcc gcatcagcaa gaccaagaag
      841 gagtacctgc tggaggagcg ggacatcaac agattctccg tgcagaagga cttcatcaac
      901 cggaatctgg tggacaccag atacgccaca cgcggcctga tgaatctgct gcggtcttat
      961 ttcagagtga acaatctgga tgtgaaggtg aagagcatca acggcggctt cacctccttt
     1021 ctgcggagaa agtggaagtt taagaaggag cgcaacaagg gctataagca ccacgccgag
     1081 gatgccctga tcatcgccaa tgccgacttc atctttaagg agtggaagaa gctggacaag
     1141 gccaagaaag tgatggagaa ccagatgttc gaggagaagc aggccgagag catgcccgag
     1201 atcgagacag agcaggagta caaggagatt ttcatcacac ctcaccagat caagcacatc
     1261 aaggacttca aggactacaa gtattctcac agggtggata agaagcccaa ccgcaagctg
     1321 atcaatgaca ccctgtatag cacacggaag gacgataagg gcaataccct gatcgtgaac
     1381 aatctgaacg gcctgtacga caaggataat gacaagctga agaagctgat caacaagtct
     1441 cccgagaagc tgctgatgta ccaccacgat cctcagacat atcagaagct gaagctgatc
     1501 atggagcagt acggcgacga gaagaaccca ctgtataagt actatgagga gacaggcaac
     1561 tacctgacaa agtatagcaa gaaggataat ggccccgtga tcaagaagat caagtactat
     1621 ggcaacaagc tgaatgccca cctggacatc accgacgatt accctaactc tcgcaataag
     1681 gtggtgaagc tgagcctgaa gccataccgg ttcgacgtgt acctggacaa cggcgtgtat
     1741 aagtttgtga cagtgaagaa tctggatgtg atcaagaagg agaactacta tgaggtgaac
     1801 agcaagtgct acgaggaggc caagaagctg aagaagatca gcaaccaggc cgagttcatc
     1861 gcctcttttt acaagaatga cctgatcaag atcaatggcg agctgtatag agtgatcggc
     1921 gtgaacaatg atctgctgaa cagaatcgaa gtgaatatga tcgacatcac ctacagggag
     1981 tatctggaga acatgaatga taagaggccc cctcatatca tcaagaccat cgcctctaag
     2041 acacagagca tcaagaagta cagcacagac atcctgggga acctgtatga agtcaagagc
     2101 aagaaacatc ctcagattat caagaaaggc agcggcggga gcggcgggag cggggggagc
     2161 actaatctga gcgacatcat tgagaaggag actgggaaac agctggtcat tcaggagtcc
     2221 atcctgatgc tgcctgagga ggtggaggaa gtgatcggca acaagccaga gtctgacatc
     2281 ctggtgcaca ccgcctacga cgagtccaca gatgagaatg tgatgctgct gacctctgac
     2341 gcccccgagt ataagccttg ggccctggtc atccaggatt ctaacggcga gaataagatc
     2401 aagatgctga gcggaggatc caaaagaacc gccgacggca gcgaattcga gcccaagaag
     2461 aagaggaaag tctaaccggt catcatcacc atcaccattg agtttaaacc cgctgatcag
     2521 cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct
     2581 tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc
     2641 attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg
     2701 aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg
     2761 cggaaagaac cagctggggc tcgataccgt cgacctctag ctagagcttg gcgtaatcat
     2821 ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag
     2881 ccggaagcat aaagtgtaaa gcctagggtg cctaatgagt gagctaactc acattaattg
     2941 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     3001 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca
     3061 ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg
     3121 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     3181 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     3241 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     3301 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     3361 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     3421 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     3481 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     3541 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     3601 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     3661 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     3721 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     3781 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     3841 ctgacactca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     3901 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     3961 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     4021 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     4081 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     4141 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     4201 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     4261 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     4321 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     4381 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     4441 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     4501 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     4561 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     4621 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     4681 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     4741 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     4801 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     4861 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     4921 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg
     4981 acggatcggg agatcgatct cccgatcccc tagggtcgac tctcagtaca atctgctctg
     5041 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
     5101 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
     5161 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
     5221 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
     5281 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
     5341 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
     5401 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
     5461 tgtatc
//
