LOCUS       Exported                7725 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  AAV genome: expresses the N-terminal of v5 AAV-ABE from the Cbh 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    Cbh_v5 AAV-ABE N-terminal
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7725)
  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 7725)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7725
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(125..142)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(125..141)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(134..156)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      283..738
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(370..389)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     580..601
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        764..868
                     /gene="bla"
                     /label=AmpR promoter
     CDS             869..1729
                     /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     complement(1087..1106)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1900..2488
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2389..2408
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2642..2659
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2776..2797
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        2812..2842
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2850..2866
                     /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)."
     primer_bind     2855..2877
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2874..2890
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2874..2890
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   2917..3046
                     /label=AAV2 ITR
     enhancer        3069..3354
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        3356..3633
                     /label=chicken beta-actin promoter
     intron          3634..3861
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     CDS             3915..3935
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             5124..6839
                     /codon_start=1
                     /product="N-terminal portion of Streptococcus pyogenes Cas9
                     (Zetsche et al., 2015)"
                     /label=Cas9(N)
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIE"
     CDS             7188..7208
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(7273..7293)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     polyA_signal    7469..7693
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   7702..106
                     /label=AAV2 ITR
ORIGIN
        1 ccactccctc tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac
       61 gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc gcgcagcctt aattaaccta
      121 attcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt acccaactta
      181 atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag gcccgcaccg
      241 atcgcccttc ccaacagttg cgcagcctga atggcgaatg ggacgcgccc tgtagcggcg
      301 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc
      361 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
      421 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
      481 accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
      541 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
      601 gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt
      661 cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa
      721 tattaacgct tacaatttag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
      781 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
      841 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
      901 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
      961 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     1021 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     1081 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     1141 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     1201 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     1261 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     1321 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     1381 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     1441 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     1501 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     1561 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     1621 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     1681 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     1741 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     1801 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     1861 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     1921 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     1981 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     2041 tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     2101 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     2161 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     2221 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     2281 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     2341 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     2401 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     2461 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     2521 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     2581 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     2641 cagcgagtca gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc ctctccccgc
     2701 gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga aagcgggcag
     2761 tgagcgcaac gcaattaatg tgagttagct cactcattag gcaccccagg ctttacactt
     2821 tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc acacaggaaa
     2881 cagctatgac catgattacg ccagatttaa ttaaggctgc gcgctcgctc gctcactgag
     2941 gccgcccggg caaagcccgg gcgtcgggcg acctttggtc gcccggcctc agtgagcgag
     3001 cgagcgcgca gagagggagt ggccaactcc atcactaggg gttcctgcgg cctctagatc
     3061 agggtacccg ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc
     3121 cgcccattga cgtcaatagt aacgccaata gggactttcc attgacgtca atgggtggag
     3181 tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc
     3241 cctattgacg tcaatgacgg taaatggccc gcctggcatt gtgcccagta catgacctta
     3301 tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac catggtcgag
     3361 gtgagcccca cgttctgctt cactctcccc atctcccccc cctccccacc cccaattttg
     3421 tatttattta ttttttaatt attttgtgca gcgatggggg cggggggggg gggggggcgc
     3481 gcgccaggcg gggcggggcg gggcgagggg cggggcgggg cgaggcggag aggtgcggcg
     3541 gcagccaatc agagcggcgc gctccgaaag tttcctttta tggcgaggcg gcggcggcgg
     3601 cggccctata aaaagcgaag cgcgcggcgg gcgggagtcg ctgcgcgctg ccttcgcccc
     3661 gtgccccgct ccgccgccgc ctcgcgccgc ccgccccggc tctgactgac cgcgttactc
     3721 ccacaggtga gcgggcggga cggcccttct cctccgggct gtaattagct gagcaagagg
     3781 taagggttta agggatggtt ggttggtggg gtattaatgt ttaattacct ggagcacctg
     3841 cctgaaatca ctttttttca ggttggaccg gtgccaccat gaaacggaca gccgacggaa
     3901 gcgagttcga gtcaccaaag aagaagcgga aagtctctga agtcgagttt agccacgagt
     3961 attggatgag gcacgcactg accctggcaa agcgagcatg ggatgaaaga gaagtccccg
     4021 tgggcgccgt gctggtgcac aacaatagag tgatcggaga gggatggaac aggccaatcg
     4081 gccgccacga ccctaccgca cacgcagaga tcatggcact gaggcaggga ggcctggtca
     4141 tgcagaatta ccgcctgatc gatgccaccc tgtatgtgac actggagcca tgcgtgatgt
     4201 gcgcaggagc aatgatccac agcaggatcg gaagagtggt gttcggagca cgggacgcca
     4261 agaccggcgc agcaggctcc ctgatggatg tgctgcacca ccccggcatg aaccaccggg
     4321 tggagatcac agagggaatc ctggcagacg agtgcgccgc cctgctgagc gatttcttta
     4381 gaatgcggag acaggagatc aaggcccaga agaaggcaca gagctccacc gactctggag
     4441 gatctagcgg aggatcctct ggaagcgaga caccaggcac aagcgagtcc gccacaccag
     4501 agagctccgg cggctcctcc ggaggatcct ctgaggtgga gttttcccac gagtactgga
     4561 tgagacatgc cctgaccctg gccaagaggg cacgcgatga gagggaggtg cctgtgggag
     4621 ccgtgctggt gctgaacaat agagtgatcg gcgagggctg gaacagagcc atcggcctgc
     4681 acgacccaac agcccatgcc gaaattatgg ccctgagaca gggcggcctg gtcatgcaga
     4741 actacagact gattgacgcc accctgtacg tgacattcga gccttgcgtg atgtgcgccg
     4801 gcgccatgat ccactctagg atcggccgcg tggtgtttgg cgtgaggaac gcaaaaaccg
     4861 gcgccgcagg ctccctgatg gacgtgctgc actaccccgg catgaatcac cgcgtcgaaa
     4921 ttaccgaggg aatcctggca gatgaatgtg ccgccctgct gtgctatttc tttcggatgc
     4981 ctagacaggt gttcaatgct cagaagaagg cccagagctc caccgactcc ggaggatcta
     5041 gcggaggctc ctctggctct gagacacctg gcacaagcga gagcgcaaca cctgaaagca
     5101 gcgggggcag cagcgggggg tcagacaaga agtacagcat cggcctggcc atcggcacca
     5161 actctgtggg ctgggccgtg atcaccgacg agtacaaggt gcccagcaag aaattcaagg
     5221 tgctgggcaa caccgaccgg cacagcatca agaagaacct gatcggagcc ctgctgttcg
     5281 acagcggcga aacagccgag gccacccggc tgaagagaac cgccagaaga agatacacca
     5341 gacggaagaa ccggatctgc tatctgcaag agatcttcag caacgagatg gccaaggtgg
     5401 acgacagctt cttccacaga ctggaagagt ccttcctggt ggaagaggat aagaagcacg
     5461 agcggcaccc catcttcggc aacatcgtgg acgaggtggc ctaccacgag aagtacccca
     5521 ccatctacca cctgagaaag aaactggtgg acagcaccga caaggccgac ctgcggctga
     5581 tctatctggc cctggcccac atgatcaagt tccggggcca cttcctgatc gagggcgacc
     5641 tgaaccccga caacagcgac gtggacaagc tgttcatcca gctggtgcag acctacaacc
     5701 agctgttcga ggaaaacccc atcaacgcca gcggcgtgga cgccaaggcc atcctgtctg
     5761 ccagactgag caagagcaga cggctggaaa atctgatcgc ccagctgccc ggcgagaaga
     5821 agaatggcct gttcggaaac ctgattgccc tgagcctggg cctgaccccc aacttcaaga
     5881 gcaacttcga cctggccgag gatgccaaac tgcagctgag caaggacacc tacgacgacg
     5941 acctggacaa cctgctggcc cagatcggcg accagtacgc cgacctgttt ctggccgcca
     6001 agaacctgtc cgacgccatc ctgctgagcg acatcctgag agtgaacacc gagatcacca
     6061 aggcccccct gagcgcctct atgatcaaga gatacgacga gcaccaccag gacctgaccc
     6121 tgctgaaagc tctcgtgcgg cagcagctgc ctgagaagta caaagagatt ttcttcgacc
     6181 agagcaagaa cggctacgcc ggctacattg acggcggagc cagccaggaa gagttctaca
     6241 agttcatcaa gcccatcctg gaaaagatgg acggcaccga ggaactgctc gtgaagctga
     6301 acagagagga cctgctgcgg aagcagcgga ccttcgacaa cggcagcatc ccccaccaga
     6361 tccacctggg agagctgcac gccattctgc ggcggcagga agatttttac ccattcctga
     6421 aggacaaccg ggaaaagatc gagaagatcc tgaccttccg catcccctac tacgtgggcc
     6481 ctctggccag gggaaacagc agattcgcct ggatgaccag aaagagcgag gaaaccatca
     6541 ccccctggaa cttcgaggaa gtggtggaca agggcgcttc cgcccagagc ttcatcgagc
     6601 ggatgaccaa cttcgataag aacctgccca acgagaaggt gctgcccaag cacagcctgc
     6661 tgtacgagta cttcaccgtg tataacgagc tgaccaaagt gaaatacgtg accgagggaa
     6721 tgagaaagcc cgccttcctg agcggcgagc agaaaaaggc catcgtggac ctgctgttca
     6781 agaccaaccg gaaagtgacc gtgaagcagc tgaaagagga ctacttcaag aaaatcgagt
     6841 gcctgtccta cgagacagag atcctgacag tggagtatgg cctgctgcca atcggcaaga
     6901 tcgtggagaa gaggatcgag tgtaccgtgt actctgtgga taacaatggc aacatctata
     6961 cacagcccgt ggcacagtgg cacgataggg gagagcagga ggtgttcgag tattgcctgg
     7021 aggacggcag cctgatcagg gcaaccaagg accacaagtt catgacagtg gatggccaga
     7081 tgctgcccat cgacgagatt ttcgagcggg agctggacct gatgagagtg gataacctgc
     7141 ctaatagcgg aggcagtaaa agaacagcag acgggagtga gtttgagccc aagaaaaaga
     7201 gaaaggtgta agatctgata atcaacctct ggattacaaa atttgtgaaa gattgactgg
     7261 tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa tgcctttgta
     7321 tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat cctggttagt
     7381 tcttgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct
     7441 gttgggcact gacaattccg tggtgcgact gtgccttcta gttgccagcc atctgttgtt
     7501 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     7561 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     7621 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg
     7681 gtgggctcta tgggcggccg caggaacccc tagtgatgga gttgg
//
