LOCUS       sequence-411180-        7137 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses TadA8e and SpG cas9N by the constitutive CMV promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7137)
  AUTHORS   Yang L, Liu Z, Sun J, Chen Z, Gao F, Guo Y
  TITLE     Adenine base editor-based correction of the cardiac pathogenic Lmna 
            c.1621C > T mutation in murine hearts.
  JOURNAL   J Cell Mol Med. 2024 Feb;28(4):e18145. doi: 10.1111/jcmm.18145.
  PUBMED    38332517
REFERENCE   2  (bases 1 to 7137)
  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: "J Cell Mol 
            Med."; date: "2024-02"; volume: "28(4):e18145. doi"; pages: " 
            10.1111/jcmm.18145"
COMMENT     Sequence Label: pAAV-CMV-TadA8e-SpG N aa2-713-InteinN
FEATURES             Location/Qualifiers
     source          1..7137
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   11..140
                     /label=AAV2 ITR
     primer_bind     complement(187..207)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     enhancer        255..634
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        635..838
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     789..809
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      866..875
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             878..898
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1493..3208
                     /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"
     primer_bind     complement(3948..3967)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     polyA_signal    3977..4058
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     4002..4021
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4197..4326
                     /label=AAV2 ITR
     primer_bind     complement(4364..4380)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4364..4380)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4377..4399)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4388..4404
                     /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(4412..4418,4419..4436,4437..4442))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4457..4478
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(4766..5354)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4846..4865)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5525..6316,6317..6385))
                     /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     6148..6167
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6386..6490)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6516..6971)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6653..6674)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6865..6884
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7098..7120
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7112..7129
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7113..7129
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 ttaattaagg ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg
       61 ggcgaccttt ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa
      121 ctccatcact aggggttcct tgtagttaat gattaacccg ccatgctact tatctactgc
      181 ggccgcggaa tgtgtgtcag ttagggtgtg gaaagtcccc aggctcccca gcaggcagaa
      241 gtatgcaaag catggacatt gattattgac tagttattaa tagtaatcaa ttacggggtc
      301 attagtccat agcccatata tggagttccg cgttacataa cttacggtaa atggcccgcc
      361 tggctgaccg cccaacgacc cccgcccatt gacgtcaata atgacgtatg ttcccatagt
      421 aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca
      481 cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg
      541 taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc ctacttggca
      601 gtacatctac gtattagtca tcgctattac catggtgatg cggttttggc agtacatcaa
      661 tgggcgtgga tagcggtttg actcacgggg atttccaagt ctccacccca ttgacgtcaa
      721 tgggagtttg ttttggcacc aaaatcaacg ggactttcca aaatgtcgta acaactccgc
      781 cccattgacg caaatgggcg gtaggcgtgt acggtgggag gtctatataa gcagagctgt
      841 accgaagccg ctagcgctac cggtcgccac catggcccca aagaagaagc ggaaagtctc
      901 tgaggtggag ttttcccacg agtactggat gagacatgcc ctgaccctgg ccaagagggc
      961 acgggatgag agggaggtgc ctgtgggagc cgtgctggtg ctgaacaata gagtgatcgg
     1021 cgagggctgg aacagagcca tcggcctgca cgacccaaca gcccatgccg aaattatggc
     1081 cctgagacag ggcggcctgg tcatgcagaa ctacagactg attgacgcca ccctgtacgt
     1141 gacattcgag ccttgcgtga tgtgcgccgg cgccatgatc cactctagga tcggccgcgt
     1201 ggtgtttggc gtgaggaact caaaaagagg cgccgcaggc tccctgatga acgtgctgaa
     1261 ctaccccggc atgaatcacc gcgtcgaaat taccgaggga atcctggcag atgaatgtgc
     1321 cgccctgctg tgcgatttct atcggatgcc tagacaggtg ttcaatgctc agaagaaggc
     1381 ccagagctcc atcaactccg gaggatctag cggaggctcc tctggctctg agacacctgg
     1441 cacaagcgag agcgcaacac ctgaaagcag cgggggcagc agcggggggt cagacaagaa
     1501 gtacagcatc ggcctggcca tcggcaccaa ctctgtgggc tgggccgtga tcaccgacga
     1561 gtacaaggtg cccagcaaga aattcaaggt gctgggcaac accgaccggc acagcatcaa
     1621 gaagaacctg atcggagccc tgctgttcga cagcggcgaa acagccgagg ccacccggct
     1681 gaagagaacc gccagaagaa gatacaccag acggaagaac cggatctgct atctgcaaga
     1741 gatcttcagc aacgagatgg ccaaggtgga cgacagcttc ttccacagac tggaagagtc
     1801 cttcctggtg gaagaggata agaagcacga gcggcacccc atcttcggca acatcgtgga
     1861 cgaggtggcc taccacgaga agtaccccac catctaccac ctgagaaaga aactggtgga
     1921 cagcaccgac aaggccgacc tgcggctgat ctatctggcc ctggcccaca tgatcaagtt
     1981 ccggggccac ttcctgatcg agggcgacct gaaccccgac aacagcgacg tggacaagct
     2041 gttcatccag ctggtgcaga cctacaacca gctgttcgag gaaaacccca tcaacgccag
     2101 cggcgtggac gccaaggcca tcctgtctgc cagactgagc aagagcagac ggctggaaaa
     2161 tctgatcgcc cagctgcccg gcgagaagaa gaatggcctg ttcggaaacc tgattgccct
     2221 gagcctgggc ctgaccccca acttcaagag caacttcgac ctggccgagg atgccaaact
     2281 gcagctgagc aaggacacct acgacgacga cctggacaac ctgctggccc agatcggcga
     2341 ccagtacgcc gacctgtttc tggccgccaa gaacctgtcc gacgccatcc tgctgagcga
     2401 catcctgaga gtgaacaccg agatcaccaa ggcccccctg agcgcctcta tgatcaagag
     2461 atacgacgag caccaccagg acctgaccct gctgaaagct ctcgtgcggc agcagctgcc
     2521 tgagaagtac aaagagattt tcttcgacca gagcaagaac ggctacgccg gctacattga
     2581 cggcggagcc agccaggaag agttctacaa gttcatcaag cccatcctgg aaaagatgga
     2641 cggcaccgag gaactgctcg tgaagctgaa cagagaggac ctgctgcgga agcagcggac
     2701 cttcgacaac ggcagcatcc cccaccagat ccacctggga gagctgcacg ccattctgcg
     2761 gcggcaggaa gatttttacc cattcctgaa ggacaaccgg gaaaagatcg agaagatcct
     2821 gaccttccgc atcccctact acgtgggccc tctggccagg ggaaacagca gattcgcctg
     2881 gatgaccaga aagagcgagg aaaccatcac cccctggaac ttcgaggaag tggtggacaa
     2941 gggcgcttcc gcccagagct tcatcgagcg gatgaccaac ttcgataaga acctgcccaa
     3001 cgagaaggtg ctgcccaagc acagcctgct gtacgagtac ttcaccgtgt ataacgagct
     3061 gaccaaagtg aaatacgtga ccgagggaat gagaaagccc gccttcctga gcggcgagca
     3121 gaaaaaggcc atcgtggacc tgctgttcaa gaccaaccgg aaagtgaccg tgaagcagct
     3181 gaaagaggac tacttcaaga aaatcgagtg cttcgactcc gtggaaatct ccggcgtgga
     3241 agatcggttc aacgcctccc tgggcacata ccacgatctg ctgaaaatta tcaaggacaa
     3301 ggacttcctg gacaatgagg aaaacgagga cattctggaa gatatcgtgc tgaccctgac
     3361 actgtttgag gacagagaga tgatcgagga acggctgaaa acctatgccc acctgttcga
     3421 cgacaaagtg atgaagcagc tgaagcggcg gagatacacc ggctggggca ggctgagccg
     3481 gaagctgatc aacggcatcc gagacaagca gagtggaaag acaatcctgg attttcttaa
     3541 gtccgatgga tttgccaacc ggaacttcat gcagttgatc catgatgact ctctcacctt
     3601 taaggaggac atccagaaag cacaagtttg tctggctggc gatactctca ttaccctggc
     3661 cgatggacga cgagtgccta ttagagaact ggtgtcacag cagaattttt ccgtgtgggc
     3721 tctgaatcct cagacttacc gcctggagag ggctagagtg agtagagctt tctgtaccgg
     3781 catcaaacct gtgtaccgcc tcaccactag actggggaga tccattaggg ccactgccaa
     3841 ccaccgattt ctcacacctc agggctggaa acgagtcgat gaactccagc ctggagatta
     3901 cctggctctg cctaggagaa tccctactgc ctcctgatct cgagctcgat gagtttggac
     3961 aaaccacaac tagaatgcag tgaaaaaaat gctttatttg tgaaatttgt gatgctattg
     4021 ctttatttgt aaccattata agctgcaata aacaagttaa caacaacaat tgcattcatt
     4081 ttatgtttca ggttcagggg gaggtgtggg aggtttttta aagcaagtaa aacctctaca
     4141 aatgtggtag cggccgcagg tagataagta gcatggcggg ttaatcatta actacaagga
     4201 acccctagtg atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg
     4261 gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc
     4321 gcgcagccta attaaggcct taattaaatc tggcgtaatc atggtcatag ctgtttcctg
     4381 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     4441 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     4501 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     4561 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     4621 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     4681 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     4741 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     4801 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     4861 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     4921 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     4981 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     5041 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     5101 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     5161 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     5221 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     5281 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     5341 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     5401 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     5461 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     5521 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     5581 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     5641 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     5701 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     5761 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     5821 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     5881 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     5941 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     6001 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     6061 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     6121 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     6181 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     6241 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     6301 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     6361 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     6421 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     6481 gggttccgcg cacatttccc cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt
     6541 aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg
     6601 caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg
     6661 gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta
     6721 tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg
     6781 ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa
     6841 gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct
     6901 ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct
     6961 acagggcgcg tcccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg
     7021 ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc gattaagttg
     7081 ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg aattagg
//
