LOCUS       sequence-272940-        8037 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A-to-G base editor.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8037)
  AUTHORS   Richter MF, Zhao KT, Eton E, Lapinaite A, Newby GA, Thuronyi BW, 
            Wilson C, Koblan LW, Zeng J, Bauer DE, Doudna JA, Liu DR
  TITLE     Phage-assisted evolution of an adenine base editor with improved Cas
            domain compatibility and activity.
  JOURNAL   Nat Biotechnol. 2020 Jul;38(7):883-891. doi: 
            10.1038/s41587-020-0453-z. Epub 2020 Mar 16.
  PUBMED    32433547
REFERENCE   2  (bases 1 to 8037)
  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/s41587-020-0453-z"; journalName: "Nat Biotechnol"; date: 
            "2020-07"; volume: "38"; issue: "7"; pages: "883-891"
COMMENT     Sequence Label: enAsABE8e
FEATURES             Location/Qualifiers
     source          1..8037
                     /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             1060..4980
                     /codon_start=1
                     /product="CRISPR-associated protein from Acidaminococcus 
                     sp. BV3L6 (Zetsche et al., 2015)"
                     /label=AsCpf1
                     /translation="MTQFEGFTNLYQVSKTLRFELIPQGKTLKHIQEQGFIEEDKARND
                     HYKELKPIIDRIYKTYADQCLQLVQLDWENLSAAIDSYRKEKTEETRNALIEEQATYRN
                     AIHDYFIGRTDNLTDAINKRHAEIYKGLFKAELFNGKVLKQLGTVTTTEHENALLRSFD
                     KFTTYFSGFYRNRKNVFSAEDISTAIPHRIVQDNFPKFKENCHIFTRLITAVPSLREHF
                     ENVKKAIGIFVSTSIEEVFSFPFYNQLLTQTQIDLYNQLLGGISREAGTEKIKGLNEVL
                     NLAIQKNDETAHIIASLPHRFIPLFKQILSDRNTLSFILEEFKSDEEVIQSFCKYKTLL
                     RNENVLETAEALFNELNSIDLTHIFISHKKLETISSALCDHWDTLRNALYERRISELTG
                     KITKSAKEKVQRSLKHEDINLQEIISAAGKELSEAFKQKTSEILSHAHAALDQPLPTTL
                     KKQEEKEILKSQLDSLLGLYHLLDWFAVDESNEVDPEFSARLTGIKLEMEPSLSFYNKA
                     RNYATKKPYSVEKFKLNFQMPTLARGWDVNREKNNGAILFVKNGLYYLGIMPKQKGRYK
                     ALSFEPTEKTSEGFDKMYYDYFPDAAKMIPKCSTQLKAVTAHFQTHTTPILLSNNFIEP
                     LEITKEIYDLNNPEKEPKKFQTAYAKKTGDQKGYREALCKWIDFTRDFLSKYTKTTSID
                     LSSLRPSSQYKDLGEYYAELNPLLYHISFQRIAEKEIMDAVETGKLYLFQIYNKDFAKG
                     HHGKPNLHTLYWTGLFSPENLAKTSIKLNGQAELFYRPKSRMKRMAHRLGEKMLNKKLK
                     DQKTPIPDTLYQELYDYVNHRLSHDLSDEARALLPNVITKEVSHEIIKDRRFTSDKFFF
                     HVPITLNYQAANSPSKFNQRVNAYLKEHPETPIIGIARGERNLIYITVIDSTGKILEQR
                     SLNTIQQFDYQKKLDNREKERVAARQAWSVVGTIKDLKQGYLSQVIHEIVDLMIHYQAV
                     VVLENLNFGFKSKRTGIAEKAVYQQFEKMLIDKLNCLVLKDYPAEKVGGVLNPYQLTDQ
                     FTSFAKMGTQSGFLFYVPAPYTSKIDPLTGFVDPFVWKTIKNHESRKHFLEGFDFLHYD
                     VKTGDFILHFKMNRNLSFQRGLPGFMPAWDIVFEKNETQFDAKGTPFIAGKRIVPVIEN
                     HRFTGRYRDLYPANELIALLEEKGIVFRDGSNILPKLLENDDSHAIDTMVALIRSVLQM
                     RNSNAATGEDYINSPVRDLNGVCFDSRFQNPEWPMDADANGAYHIALKGQLLLNHLKES
                     KDLKLQNGISNQDWLAYIQELRN"
     regulatory      4700..4709
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             5023..5043
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             5052..5069
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(5092..5109)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5098..5322
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5393..5409)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5393..5409)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5406..5428)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5417..5433
                     /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(5441..5447,5448..5465,5466..5471))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5486..5507
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5624..5641)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5795..6383)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5875..5894)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6554..7345,7346..7414))
                     /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     7177..7196
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7415..7519)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7598..7617)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        join(7789..8037,1..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 aagtctctga ggtggagttt
      481 tcccacgagt actggatgag acatgccctg accctggcca agagggcacg ggatgagagg
      541 gaggtgcctg tgggagccgt gctggtgctg aacaatagag tgatcggcga gggctggaac
      601 agagccatcg gcctgcacga cccaacagcc catgccgaaa ttatggccct gagacagggc
      661 ggcctggtca tgcagaacta cagactgatt gacgccaccc tgtacgtgac attcgagcct
      721 tgcgtgatgt gcgccggcgc catgatccac tctaggatcg gccgcgtggt gtttggcgtg
      781 aggaactcaa aaagaggcgc cgcaggctcc ctgatgaacg tgctgaacta ccccggcatg
      841 aatcaccgcg tcgaaattac cgagggaatc ctggcagatg aatgtgccgc cctgctgtgc
      901 gatttctatc ggatgcctag acaggtgttc aatgctcaga agaaggccca gagctccatc
      961 aactccggag gatctagcgg aggatcctct ggcagcgaga caccaggaac aagcgagtca
     1021 gcaacaccag agagcagtgg cggcagcagc ggggggtcaa tgacacagtt cgagggcttt
     1081 accaacctgt atcaggtgag caagacactg cggtttgagc tgatcccaca gggcaagacc
     1141 ctgaagcaca tccaggagca gggcttcatc gaggaggaca aggcccgcaa tgatcactac
     1201 aaggagctga agcccatcat cgatcggatc tacaagacct atgccgacca gtgcctgcag
     1261 ctggtgcagc tggattggga gaacctgagc gccgccatcg actcctatag aaaggagaaa
     1321 accgaggaga caaggaacgc cctgatcgag gagcaggcca catatcgcaa tgccatccac
     1381 gactacttca tcggccggac agacaacctg accgatgcca tcaataagag acacgccgag
     1441 atctacaagg gcctgttcaa ggccgagctg tttaatggca aggtgctgaa gcagctgggc
     1501 accgtgacca caaccgagca cgagaacgcc ctgctgcgga gcttcgacaa gtttacaacc
     1561 tacttctccg gcttttatag aaacaggaag aacgtgttca gcgccgagga tatcagcaca
     1621 gccatcccac accgcatcgt gcaggacaac ttccccaagt ttaaggagaa ttgtcacatc
     1681 ttcacacgcc tgatcaccgc cgtgcccagc ctgcgggagc actttgagaa cgtgaagaag
     1741 gccatcggca tcttcgtgag cacctccatc gaggaggtgt tttccttccc tttttataac
     1801 cagctgctga cacagaccca gatcgacctg tataaccagc tgctgggagg aatctctcgg
     1861 gaggcaggca ccgagaagat caagggcctg aacgaggtgc tgaatctggc catccagaag
     1921 aatgatgaga cagcccacat catcgcctcc ctgccacaca gattcatccc cctgtttaag
     1981 cagatcctgt ccgataggaa caccctgtct ttcatcctgg aggagtttaa gagcgacgag
     2041 gaagtgatcc agtccttctg caagtacaag acactgctga gaaacgagaa cgtgctggag
     2101 acagccgagg ccctgtttaa cgagctgaac agcatcgacc tgacacacat cttcatcagc
     2161 cacaagaagc tggagacaat cagcagcgcc ctgtgcgacc actgggatac actgaggaat
     2221 gccctgtatg agcggagaat ctccgagctg acaggcaaga tcaccaagtc tgccaaggag
     2281 aaggtgcagc gcagcctgaa gcacgaggat atcaacctgc aggagatcat ctctgccgca
     2341 ggcaaggagc tgagcgaggc cttcaagcag aaaaccagcg agatcctgtc ccacgcacac
     2401 gccgccctgg atcagccact gcctacaacc ctgaagaagc aggaggagaa ggagatcctg
     2461 aagtctcagc tggacagcct gctgggcctg taccacctgc tggactggtt tgccgtggat
     2521 gagtccaacg aggtggaccc cgagttctct gcccggctga ccggcatcaa gctggagatg
     2581 gagccttctc tgagcttcta caacaaggcc agaaattatg ccaccaagaa gccctactcc
     2641 gtggagaagt tcaagctgaa ctttcagatg cctacactgg ccagaggctg ggacgtgaat
     2701 agagagaaga acaatggcgc catcctgttt gtgaagaacg gcctgtacta tctgggcatc
     2761 atgccaaagc agaagggcag gtataaggcc ctgagcttcg agcccacaga gaaaaccagc
     2821 gagggctttg ataagatgta ctatgactac ttccctgatg ccgccaagat gatcccaaag
     2881 tgcagcaccc agctgaaggc cgtgacagcc cactttcaga cccacacaac ccccatcctg
     2941 ctgtccaaca atttcatcga gcctctggag atcacaaagg agatctacga cctgaacaat
     3001 cctgagaagg agccaaagaa gtttcagaca gcctacgcca agaaaaccgg cgaccagaag
     3061 ggctacagag aggccctgtg caagtggatc gacttcacaa gggattttct gtccaagtat
     3121 accaagacaa cctctatcga tctgtctagc ctgcggccat cctctcagta taaggacctg
     3181 ggcgagtact atgccgagct gaatcccctg ctgtaccaca tcagcttcca gagaatcgcc
     3241 gagaaggaga tcatggatgc cgtggagaca ggcaagctgt acctgttcca gatctataac
     3301 aaggactttg ccaagggcca ccacggcaag cctaatctgc acacactgta ttggaccggc
     3361 ctgttttctc cagagaacct ggccaagaca agcatcaagc tgaatggcca ggccgagctg
     3421 ttctaccgcc ctaagtccag gatgaagagg atggcacacc ggctgggaga gaagatgctg
     3481 aacaagaagc tgaaggatca gaaaacccca atccccgaca ccctgtacca ggagctgtac
     3541 gactatgtga atcacagact gtcccacgac ctgtctgatg aggccagggc cctgctgccc
     3601 aacgtgatca ccaaggaggt gtctcacgag atcatcaagg ataggcgctt taccagcgac
     3661 aagttctttt tccacgtgcc tatcacactg aactatcagg ccgccaattc cccatctaag
     3721 ttcaaccaga gggtgaatgc ctacctgaag gagcaccccg agacacctat catcggcatc
     3781 gcccggggtg agagaaacct gatctatatc acagtgatcg actccaccgg caagatcctg
     3841 gagcagcgga gcctgaacac catccagcag tttgattacc agaagaagct ggacaacagg
     3901 gagaaggaga gggtggcagc aaggcaggcc tggtctgtgg tgggcacaat caaggatctg
     3961 aagcagggct atctgagcca ggtcatccac gagatcgtgg acctgatgat ccactaccag
     4021 gccgtggtgg tgctggagaa cctgaatttc ggctttaaga gcaagaggac cggcatcgcc
     4081 gagaaggccg tgtaccagca gttcgagaag atgctgatcg ataagctgaa ttgcctggtg
     4141 ctgaaggact atccagcaga gaaagtggga ggcgtgctga acccatacca gctgacagac
     4201 cagttcacct cctttgccaa gatgggcacc cagtctggct tcctgtttta cgtgcctgcc
     4261 ccatatacat ctaagatcga tcccctgacc ggcttcgtgg accccttcgt gtggaaaacc
     4321 atcaagaatc acgagagccg caagcacttc ctggagggct tcgactttct gcactacgac
     4381 gtgaaaaccg gcgacttcat cctgcacttt aagatgaaca gaaatctgtc cttccagagg
     4441 ggcctgcccg gctttatgcc tgcatgggat atcgtgttcg agaagaacga gacacagttt
     4501 gacgccaagg gcaccccttt catcgccggc aagagaatcg tgccagtgat cgagaatcac
     4561 agattcaccg gcagataccg ggacctgtat cctgccaacg agctgatcgc cctgctggag
     4621 gagaagggca tcgtgttcag ggatggctcc aacatcctgc caaagctgct ggagaatgac
     4681 gattctcacg ccatcgacac catggtggcc ctgatccgca gcgtgctgca gatgcggaac
     4741 tccaatgccg ccacaggcga ggactatatc aacagccccg tgcgcgatct gaatggcgtg
     4801 tgcttcgact cccggtttca gaacccagag tggcccatgg acgccgatgc caatggcgcc
     4861 taccacatcg ccctgaaggg ccagctgctg ctgaatcacc tgaaggagag caaggatctg
     4921 aagctgcaga acggcatctc caatcaggac tggctggcct acatccagga gctgcgcaac
     4981 tctggcggct caaaaagaac cgccgacggc agcgaattcg agcccaagaa gaagaggaaa
     5041 gtctaaccgg tcatcatcac catcaccatt gagtttaaac ccgctgatca gcctcgactg
     5101 tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg
     5161 aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga
     5221 gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg
     5281 aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa
     5341 ccagctgggg ctcgataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc
     5401 tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca
     5461 taaagtgtaa agcctagggt gcctaatgag tgagctaact cacattaatt gcgttgcgct
     5521 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     5581 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     5641 tgcgctcggt cgttcggctg cagcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     5701 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     5761 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     5821 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     5881 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     5941 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     6001 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     6061 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     6121 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     6181 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
     6241 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     6301 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     6361 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacactc
     6421 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     6481 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     6541 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     6601 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     6661 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     6721 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     6781 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     6841 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     6901 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     6961 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     7021 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     7081 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     7141 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     7201 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     7261 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     7321 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     7381 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     7441 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     7501 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg
     7561 gagatcgatc tcccgatccc ctagggtcga ctctcagtac aatctgctct gatgccgcat
     7621 agttaagcca gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca
     7681 aaatttaagc tacaacaagg caaggcttga ccgacaattg catgaagaat ctgcttaggg
     7741 ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgcgttga cattgattat
     7801 tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatattgagt
     7861 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
     7921 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
     7981 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatc
//
