LOCUS       sequence-454013-        7873 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encoding SaKKHABE8e and sgRNA cassette on a single AAV.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7873)
  AUTHORS   Liu Z, Yang L, Yang Y, Li J, Chen Z, Guo C, Guo Q, Li Q, Zhao D, Hu 
            X, Gao F, Guo Y
  TITLE     ABE-Mediated Cardiac Gene Silencing via Single AAVs Requires DNA 
            Accessibility.
  JOURNAL   Circ Res. 2025 Jan 16. doi: 10.1161/CIRCRESAHA.124.325611.
  PUBMED    39817340
REFERENCE   2  (bases 1 to 7873)
  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: "Circ Res. 
            2025 Jan 16. doi: 10.1161/CIRCRESAHA.124.325611."
COMMENT     Sequence Label: pAAV-EFS-TadA8e-SaKKH-U6-sgRNA scaffold
FEATURES             Location/Qualifiers
     source          1..7873
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     primer_bind     complement(149..166)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(149..165)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(158..180)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      307..762
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(394..413)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     604..625
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        788..892
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(893..961,962..1753)
                     /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(1111..1130)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1924..2512
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2413..2432
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    2800..2821
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2836..2841,2842..2859,2860..2866)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2874..2890
                     /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     2879..2901
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2898..2914
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2898..2914
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   2941..3070
                     /label=AAV2 ITR
     promoter        3136..3347
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             3397..3417
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7213..7233
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    7247..7471
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        complement(7602..7850)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(7660..7679)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(7830..7850)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
ORIGIN
        1 aggaacccct agtgatggag ttggccactc cctctctgcg cgctcgctcg ctcactgagg
       61 ccgggcgacc aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc
      121 gagcgcgcag ccttaattaa cctaattcac tggccgtcgt tttacaacgt cgtgactggg
      181 aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc
      241 gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg
      301 aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
      361 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
      421 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
      481 gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta
      541 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
      601 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg
      661 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
      721 aatttaacgc gaattttaac aaaatattaa cgcttacaat ttaggtggca cttttcgggg
      781 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
      841 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
      901 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
      961 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
     1021 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     1081 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     1141 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     1201 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     1261 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
     1321 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     1381 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     1441 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
     1501 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     1561 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     1621 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     1681 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     1741 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
     1801 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     1861 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     1921 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     1981 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     2041 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     2101 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     2161 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     2221 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     2281 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     2341 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     2401 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     2461 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     2521 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     2581 tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc
     2641 tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc
     2701 aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag
     2761 gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca
     2821 ttaggcaccc caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag
     2881 cggataacaa tttcacacag gaaacagcta tgaccatgat tacgccagat ttaattaagg
     2941 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
     3001 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
     3061 aggggttcct gcggcctcta gaattcgcta gctaggtctt gaaaggagtg ggaattggct
     3121 ccggtgcccg tcagtgggca gagcgcacat cgcccacagt ccccgagaag ttggggggag
     3181 gggtcggcaa ttgatccggt gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg
     3241 tcgtgtactg gctccgcctt tttcccgagg gtgggggaga accgtatata agtgcagtag
     3301 tcgccgtgaa cgttcttttt cgcaacgggt ttgccgccag aacacaggac cggtgccacc
     3361 atgaaacgga cagccgacgg aagcgagttc gagtcaccaa agaagaagcg gaaagtctct
     3421 gaggtggagt tttcccacga gtactggatg agacatgccc tgaccctggc caagagggca
     3481 cgggatgaga gggaggtgcc tgtgggagcc gtgctggtgc tgaacaatag agtgatcggc
     3541 gagggctgga acagagccat cggcctgcac gacccaacag cccatgccga aattatggcc
     3601 ctgagacagg gcggcctggt catgcagaac tacagactga ttgacgccac cctgtacgtg
     3661 acattcgagc cttgcgtgat gtgcgccggc gccatgatcc actctaggat cggccgcgtg
     3721 gtgtttggcg tgaggaactc aaaaagaggc gccgcaggct ccctgatgaa cgtgctgaac
     3781 taccccggca tgaatcaccg cgtcgaaatt accgagggaa tcctggcaga tgaatgtgcc
     3841 gccctgctgt gcgatttcta tcggatgcct agacaggtgt tcaatgctca gaagaaggcc
     3901 cagagctcca tcaactccgg aggatctagc ggaggctcct ctggctctga gacacctggc
     3961 acaagcgaga gcgcaacacc tgaaagcagc gggggcagca gcggggggtc agggaagcga
     4021 aattacattc tggggctggc cattggcatt acatcagtgg gctatggcat cattgactac
     4081 gagacaaggg acgtgatcga cgccggcgtg agactgttca aggaggccaa cgtggagaac
     4141 aatgagggcc ggagatccaa gaggggagca aggcgcctga agcggagaag gcgccacaga
     4201 atccagagag tgaagaagct gctgttcgat tacaacctgc tgaccgacca ctccgagctg
     4261 tctggcatca atccttatga ggccagagtg aagggcctgt cccagaagct gtctgaggag
     4321 gagtttagcg ccgccctgct gcacctggca aagaggagag gcgtgcacaa cgtgaatgag
     4381 gtggaggagg acaccggcaa cgagctgtcc acaaaggagc agatcagccg caattccaag
     4441 gccctggagg agaagtatgt ggccgagctg cagctggagc ggctgaagaa ggatggcgag
     4501 gtgaggggct ccatcaatcg cttcaagacc tctgactacg tgaaggaggc caagcagctg
     4561 ctgaaggtgc agaaggccta ccaccagctg gatcagtcct ttatcgatac atatatcgac
     4621 ctgctggaga caaggcgcac atactatgag ggaccaggag agggctctcc cttcggctgg
     4681 aaggacatca aggagtggta cgagatgctg atgggccact gcacctattt tccagaggag
     4741 ctgagaagcg tgaagtacgc ctataacgcc gatctgtaca acgccctgaa tgacctgaac
     4801 aacctggtca tcaccaggga tgagaacgag aagctggagt actatgagaa gttccagatc
     4861 atcgagaacg tgttcaagca gaagaagaag cctacactga agcagatcgc caaggagatc
     4921 ctggtgaacg aggaggacat caagggctac cgcgtgacct ccacaggcaa gccagagttc
     4981 accaatctga aggtgtatca cgatatcaag gacatcacag cccggaagga gatcatcgag
     5041 aacgccgagc tgctggatca gatcgccaag atcctgacca tctatcagag ctccgaggac
     5101 atccaggagg agctgaccaa cctgaatagc gagctgacac aggaggagat cgagcagatc
     5161 agcaatctga agggctacac cggcacacac aacctgagcc tgaaggccat caatctgatc
     5221 ctggatgagc tgtggcacac aaacgacaat cagatcgcca tctttaaccg gctgaagctg
     5281 gtgccaaaga aggtggacct gtcccagcag aaggagatcc caaccacact ggtggacgat
     5341 ttcatcctgt ctcccgtggt gaagcggagc ttcatccaga gcatcaaagt gatcaacgcc
     5401 atcatcaaga agtacggcct gcccaatgat atcatcatcg agctggccag ggagaagaac
     5461 tccaaggacg cccagaagat gatcaatgag atgcagaaga ggaaccgcca gaccaatgag
     5521 cggatcgagg agatcatcag aaccacaggc aaggagaacg ccaagtacct gatcgagaag
     5581 atcaagctgc acgatatgca ggagggcaag tgtctgtatt ctctggaggc catccctctg
     5641 gaggacctgc tgaacaatcc attcaactac gaggtggatc acatcatccc ccggagcgtg
     5701 agcttcgaca attcttttaa caataaggtg ctggtgaagc aggaggagaa cagcaagaag
     5761 ggcaatagga cccctttcca gtacctgtct agctccgatt ctaagatcag ctacgagaca
     5821 ttcaagaagc acatcctgaa tctggccaag ggcaagggcc gcatcagcaa gaccaagaag
     5881 gagtacctgc tggaggagcg ggacatcaac agattctccg tgcagaagga cttcatcaac
     5941 cggaatctgg tggacaccag atacgccaca cgcggcctga tgaatctgct gcggtcttat
     6001 ttcagagtga acaatctgga tgtgaaggtg aagagcatca acggcggctt cacctccttt
     6061 ctgcggagaa agtggaagtt taagaaggag cgcaacaagg gctataagca ccacgccgag
     6121 gatgccctga tcatcgccaa tgccgacttc atctttaagg agtggaagaa gctggacaag
     6181 gccaagaaag tgatggagaa ccagatgttc gaggagaagc aggccgagag catgcccgag
     6241 atcgagacag agcaggagta caaggagatt ttcatcacac ctcaccagat caagcacatc
     6301 aaggacttca aggactacaa gtattctcac agggtggata agaagcccaa ccgcaagctg
     6361 atcaatgaca ccctgtatag cacacggaag gacgataagg gcaataccct gatcgtgaac
     6421 aatctgaacg gcctgtacga caaggataat gacaagctga agaagctgat caacaagtct
     6481 cccgagaagc tgctgatgta ccaccacgat cctcagacat atcagaagct gaagctgatc
     6541 atggagcagt acggcgacga gaagaaccca ctgtataagt actatgagga gacaggcaac
     6601 tacctgacaa agtatagcaa gaaggataat ggccccgtga tcaagaagat caagtactat
     6661 ggcaacaagc tgaatgccca cctggacatc accgacgatt accctaactc tcgcaataag
     6721 gtggtgaagc tgagcctgaa gccataccgg ttcgacgtgt acctggacaa cggcgtgtat
     6781 aagtttgtga cagtgaagaa tctggatgtg atcaagaagg agaactacta tgaggtgaac
     6841 agcaagtgct acgaggaggc caagaagctg aagaagatca gcaaccaggc cgagttcatc
     6901 gcctcttttt acaagaatga cctgatcaag atcaatggcg agctgtatag agtgatcggc
     6961 gtgaacaatg atctgctgaa cagaatcgaa gtgaatatga tcgacatcac ctacagggag
     7021 tatctggaga acatgaatga taagaggccc cctcatatca tcaagaccat cgcctctaag
     7081 acacagagca tcaagaagta cagcacagac atcctgggga acctgtatga agtcaagagc
     7141 aagaaacatc ctcagattat caagaaaggc tctggcggct caaaaagaac cgccgacggc
     7201 agcgaattcg agcccaagaa gaagaggaaa gtctaataga tctcgactgt gccttctagt
     7261 tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact
     7321 cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat
     7381 tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga agacaatagc
     7441 aggcatgctg gggatgcggt gggctctatg gctcgagcgg cccaagctta aaaaaatctc
     7501 gccaacaagt tgacgagata aacacggcat tttgccttgt tttagtagat tctgtaattt
     7561 tcattacaga gtactaaaac cgagacgcat taatcgtctc gggtgtttcg tcctttccac
     7621 aagatatata aagccaagaa atcgaaatac tttcaagtta cggtaagcat atgatagtcc
     7681 attttaaaac ataattttaa aactgcaaac tacccaagaa attattactt tctacgtcac
     7741 gtattttgta ctaatatctt tgtgtttaca gtcaaattaa ttctaattat ctctctaaca
     7801 gccttgtatc gtatatgcaa atatgaagga atcatgggaa ataggccctc ttcctgcccg
     7861 accttgcggc cgc
//
