LOCUS       sequence-410370-        8217 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Expresses ABE8r in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8217)
  AUTHORS   Xiao YL, Wu Y, Tang W
  TITLE     An adenine base editor variant expands context compatibility.
  JOURNAL   Nat Biotechnol. 2024 Jan 2. doi: 10.1038/s41587-023-01994-3.
  PUBMED    38168987
REFERENCE   2  (bases 1 to 8217)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Biotechnol. 2024 Jan 2. doi: 10.1038/s41587-023-01994-3."
COMMENT     Sequence Label: ABE8r
FEATURES             Location/Qualifiers
     source          1..8217
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(5..22)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    11..235
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(306..322)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(306..322)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(319..341)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    330..346
                     /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(354..360,361..378,379..384))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    399..420
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(537..554)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(708..1296)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(788..807)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1467..2258,2259..2327))
                     /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     2090..2109
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2328..2432)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(2511..2530)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        2702..3081
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3082..3285
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3236..3256
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     3327..3346
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3327..3345
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             3395..3415
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(4010..4033,4034..4036,4037..8110)
                     /codon_start=1
                     /product="nickase mutant of the Cas9 endonuclease from the 
                     Streptococcus pyogenes Type II CRISPR/Cas system"
                     /label=Cas9(D10A)
                     /note="generates RNA-guided single strand nicks in DNA due 
                     to the D10A mutation in the RuvC catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             8153..8173
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8182..8199
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
ORIGIN
        1 tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct
       61 tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca
      121 tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag
      181 ggggaggatt gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct
      241 gaggcggaaa gaaccagctg gggctcgata ccgtcgacct ctagctagag cttggcgtaa
      301 tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata
      361 cgagccggaa gcataaagtg taaagcctag ggtgcctaat gagtgagcta actcacatta
      421 attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa
      481 tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg
      541 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
      601 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
      661 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
      721 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
      781 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
      841 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
      901 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
      961 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     1021 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     1081 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     1141 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     1201 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     1261 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     1321 gggtctgaca ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     1381 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     1441 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     1501 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     1561 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcggct accacgctca
     1621 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     1681 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     1741 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     1801 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     1861 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     1921 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     1981 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     2041 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     2101 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     2161 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     2221 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     2281 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     2341 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     2401 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac
     2461 gtcgacggat cgggagatcg atctcccgat cccctagggt cgactctcag tacaatctgc
     2521 tctgatgccg catagttaag ccagtatctg ctccctgctt gtgtgttgga ggtcgctgag
     2581 tagtgcgcga gcaaaattta agctacaaca aggcaaggct tgaccgacaa ttgcatgaag
     2641 aatctgctta gggttaggcg ttttgcgctg cttcgcgatg tacgggccag atatacgcgt
     2701 tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt agttcatagc
     2761 ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc
     2821 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg
     2881 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat
     2941 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc
     3001 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta
     3061 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag
     3121 cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt
     3181 tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa
     3241 atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctggttt agtgaaccgt
     3301 cagatccgct agagatccgc ggccgctaat acgactcact atagggagag ccgccaccat
     3361 gaaacggaca gccgacggaa gcgagttcga gtcaccaaag aagaagcgga aagtcagcga
     3421 ggttgaattc agccacgagt actggatgcg gcatgccctg accctggcca agcgggcccg
     3481 ggatgagaga gaagtgcctg tgggcgctgt gctggtcctg aacaaccggg tgatcggcga
     3541 gggctggaac aaggccatcg gcctgcacga ccccacagcc cacgccgaga tcatggccct
     3601 gagacagggc ggtctggtga tgcagaatta tagactgtac gacgcgaccc tgtacagcac
     3661 actggaacct tgtgtgatgt gcgccggagc tatgatccac agcagaatcg gaagagtggt
     3721 gttcggcgtg cggggcgcca gacacggcgc cgtcggctct ctgatgaacg tgctccacta
     3781 cccaggcatc aagcacagag tggaaatcac cgagggaatt ctggctgatg agtgcgccgc
     3841 tctgctgtgc agattcttca ggatgcctcg cagagtgttt aaggcccaga aaaaggccca
     3901 atccagcacc gactctggag gatctagcgg aggatcctct ggcagcgaga caccaggaac
     3961 aagcgagtca gcaacaccag agagcagtgg cggcagcagc ggcggcagcg acaagaagta
     4021 cagcatcggc ctggccatcg gcaccaactc tgtgggctgg gccgtgatca ccgacgagta
     4081 caaggtgccc agcaagaaat tcaaggtgct gggcaacacc gaccggcaca gcatcaagaa
     4141 gaacctgatc ggagccctgc tgttcgacag cggcgaaaca gccgaggcca cccggctgaa
     4201 gagaaccgcc agaagaagat acaccagacg gaagaaccgg atctgctatc tgcaagagat
     4261 cttcagcaac gagatggcca aggtggacga cagcttcttc cacagactgg aagagtcctt
     4321 cctggtggaa gaggataaga agcacgagcg gcaccccatc ttcggcaaca tcgtggacga
     4381 ggtggcctac cacgagaagt accccaccat ctaccacctg agaaagaaac tggtggacag
     4441 caccgacaag gccgacctgc ggctgatcta tctggccctg gcccacatga tcaagttccg
     4501 gggccacttc ctgatcgagg gcgacctgaa ccccgacaac agcgacgtgg acaagctgtt
     4561 catccagctg gtgcagacct acaaccagct gttcgaggaa aaccccatca acgccagcgg
     4621 cgtggacgcc aaggccatcc tgtctgccag actgagcaag agcagacggc tggaaaatct
     4681 gatcgcccag ctgcccggcg aaaagaagaa tggcctgttc ggaaacctga ttgccctgag
     4741 cctgggcctg acccccaact tcaagagcaa cttcgacctg gccgaggatg ccaaactgca
     4801 gctgagcaag gacacctacg acgacgacct ggacaacctg ctggcccaga tcggcgacca
     4861 gtacgccgac ctgtttctgg ccgccaagaa cctgtccgac gccatcctgc tgagcgacat
     4921 cctgagagtg aacaccgaga tcaccaaggc ccccctgagc gcctctatga tcaagagata
     4981 cgacgagcac caccaggacc tgaccctgct gaaagctctc gtgcggcagc agctgcctga
     5041 gaagtacaaa gagattttct tcgaccagag caagaacggc tacgccggct acattgacgg
     5101 cggagccagc caggaagagt tctacaagtt catcaagccc atcctggaaa agatggacgg
     5161 caccgaggaa ctgctcgtga agctgaacag agaggacctg ctgcggaagc agcggacctt
     5221 cgacaacggc agcatccccc accagatcca cctgggagag ctgcacgcca ttctgcggcg
     5281 gcaggaagat ttttacccat tcctgaagga caaccgggaa aagatcgaga agatcctgac
     5341 cttccgcatc ccctactacg tgggccctct ggccagggga aacagcagat tcgcctggat
     5401 gaccagaaag agcgaggaaa ccatcacccc ctggaacttc gaggaagtgg tggacaaggg
     5461 cgcttccgcc cagagcttca tcgagcggat gaccaacttc gataagaacc tgcccaacga
     5521 gaaggtgctg cccaagcaca gcctgctgta cgagtacttc accgtgtata acgagctgac
     5581 caaagtgaaa tacgtgaccg agggaatgag aaagcccgcc ttcctgagcg gcgagcagaa
     5641 aaaggccatc gtggacctgc tgttcaagac caaccggaaa gtgaccgtga agcagctgaa
     5701 agaggactac ttcaagaaaa tcgagtgctt cgactccgtg gaaatctccg gcgtggaaga
     5761 tcggttcaac gcctccctgg gcacatacca cgatctgctg aaaattatca aggacaagga
     5821 cttcctggac aatgaggaaa acgaggacat tctggaagat atcgtgctga ccctgacact
     5881 gtttgaggac agagagatga tcgaggaacg gctgaaaacc tatgcccacc tgttcgacga
     5941 caaagtgatg aagcagctga agcggcggag atacaccggc tggggcaggc tgagccggaa
     6001 gctgatcaac ggcatccggg acaagcagtc cggcaagaca atcctggatt tcctgaagtc
     6061 cgacggcttc gccaacagaa acttcatgca gctgatccac gacgacagcc tgacctttaa
     6121 agaggacatc cagaaagccc aggtgtccgg ccagggcgat agcctgcacg agcacattgc
     6181 caatctggcc ggcagccccg ccattaagaa gggcatcctg cagacagtga aggtggtgga
     6241 cgagctcgtg aaagtgatgg gccggcacaa gcccgagaac atcgtgatcg aaatggccag
     6301 agagaaccag accacccaga agggacagaa gaacagccgc gagagaatga agcggatcga
     6361 agagggcatc aaagagctgg gcagccagat cctgaaagaa caccccgtgg aaaacaccca
     6421 gctgcagaac gagaagctgt acctgtacta cctgcagaat gggcgggata tgtacgtgga
     6481 ccaggaactg gacatcaacc ggctgtccga ctacgatgtg gaccatatcg tgcctcagag
     6541 ctttctgaag gacgactcca tcgacaacaa ggtgctgacc agaagcgaca agaaccgggg
     6601 caagagcgac aacgtgccct ccgaagaggt cgtgaagaag atgaagaact actggcggca
     6661 gctgctgaac gccaagctga ttacccagag aaagttcgac aatctgacca aggccgagag
     6721 aggcggcctg agcgaactgg ataaggccgg cttcatcaag agacagctgg tggaaacccg
     6781 gcagatcaca aagcacgtgg cacagatcct ggactcccgg atgaacacta agtacgacga
     6841 gaatgacaag ctgatccggg aagtgaaagt gatcaccctg aagtccaagc tggtgtccga
     6901 tttccggaag gatttccagt tttacaaagt gcgcgagatc aacaactacc accacgccca
     6961 cgacgcctac ctgaacgccg tcgtgggaac cgccctgatc aaaaagtacc ctaagctgga
     7021 aagcgagttc gtgtacggcg actacaaggt gtacgacgtg cggaagatga tcgccaagag
     7081 cgagcaggaa atcggcaagg ctaccgccaa gtacttcttc tacagcaaca tcatgaactt
     7141 tttcaagacc gagattaccc tggccaacgg cgagatccgg aagcggcctc tgatcgagac
     7201 aaacggcgaa accggggaga tcgtgtggga taagggccgg gattttgcca ccgtgcggaa
     7261 agtgctgagc atgccccaag tgaatatcgt gaaaaagacc gaggtgcaga caggcggctt
     7321 cagcaaagag tctatcctgc ccaagaggaa cagcgataag ctgatcgcca gaaagaagga
     7381 ctgggaccct aagaagtacg gcggcttcga cagccccacc gtggcctatt ctgtgctggt
     7441 ggtggccaaa gtggaaaagg gcaagtccaa gaaactgaag agtgtgaaag agctgctggg
     7501 gatcaccatc atggaaagaa gcagcttcga gaagaatccc atcgactttc tggaagccaa
     7561 gggctacaaa gaagtgaaaa aggacctgat catcaagctg cctaagtact ccctgttcga
     7621 gctggaaaac ggccggaaga gaatgctggc ctctgccggc gaactgcaga agggaaacga
     7681 actggccctg ccctccaaat atgtgaactt cctgtacctg gccagccact atgagaagct
     7741 gaagggctcc cccgaggata atgagcagaa acagctgttt gtggaacagc acaagcacta
     7801 cctggacgag atcatcgagc agatcagcga gttctccaag agagtgatcc tggccgacgc
     7861 taatctggac aaagtgctgt ccgcctacaa caagcaccgg gataagccca tcagagagca
     7921 ggccgagaat atcatccacc tgtttaccct gaccaatctg ggagcccctg ccgccttcaa
     7981 gtactttgac accaccatcg accggaagag gtacaccagc accaaagagg tgctggacgc
     8041 caccctgatc caccagagca tcaccggcct gtacgagaca cggatcgacc tgtctcagct
     8101 gggaggtgac tctggcggct caaaaagaac cgccgacggc agcgaattcg agcccaagaa
     8161 gaagaggaaa gtctaaccgg tcatcatcac catcaccatt gagtttaaac ccgctga
//
