LOCUS       sequence-436803-        6544 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Sauri cas9C by MYL2 promoter and sgRNA targeting murine 
            Camk2d intron7 5' splice site by U6 promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6544)
  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 6544)
  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-MYL2-inteinC-Sauri C aa439-1061-U6-Camk2d...
FEATURES             Location/Qualifiers
     source          1..6544
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   12..141
                     /label=AAV2 ITR
     regulatory      559..568
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      563..572
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2632..2652
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    2666..2890
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        complement(3022..3270)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(3080..3099)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(3250..3270)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     repeat_region   3605..3734
                     /label=AAV2 ITR
     primer_bind     complement(3753..3770)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3753..3769)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3762..3784)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3911..4366
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3998..4017)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4208..4229
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4392..4496
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4497..4565,4566..5357)
                     /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(4715..4734)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5528..6116
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6017..6036
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    6404..6425
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(6440..6445,6446..6463,6464..6470)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6478..6494
                     /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     6483..6505
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     6502..6518
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6502..6518
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ccttaattag gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc ttgtagttaa tgattaaccc gccatgctac ttatctacgt
      181 agccatgctc taggaagatc ggaattcgga gttccggggt tactcttccc cattagacaa
      241 tggcagggaa gagagcacac cccatcatcc ccaggccagg ccccagccac tgactcttta
      301 accttgaagg catttttggg tctcacgtgt ccacccaggc gggtggccgc ctttgagcag
      361 ctcttacttc agaagaacgg catggagtgg ggggtggggg gcttaggtgg cctccgcctc
      421 acctacaact gccaaaagtg gtcatggggt tatttttaac cccaggggag aggtatttat
      481 tgttccacag caggggcaga ggccagcagg ctcctcgaac tctccagagg tggcaactgg
      541 cctcagacac cgaccggtgc caccatggcg gcggcgtgcc cggaactgcg tcagctggcg
      601 cagagcgatg tgtattggga tccgattgtg agcattgaac cggatggcgt ggaagaagtg
      661 tttgatctga ccgtgccggg cccgcataac tttgtggcga acgatattat tgcgcataac
      721 agcgagatcg acagcatccc taccacactg gtggatgagt tcatcctgag ccctgtggtg
      781 aagcgggcct tcatccagtc catcaaggtg atcaacgctg tgatcaacag attcggcctg
      841 cccgaggaca tcattatcga gctggccaga gagaagaaca gcaaggacag aaggaagttc
      901 atcaacaagc tgcagaaaca gaatgaggcc acccggaaaa aaatcgagca gctgctggcc
      961 aagtacggca ataccaatgc caagtacatg atcgaaaaga tcaagctgca tgacatgcag
     1021 gagggcaagt gtctgtacag cctggaagct atccccctgg aagacctgct gtctaatcct
     1081 acacactacg aggtggacca catcatccct agaagcgtgt ccttcgacaa cagcctgaac
     1141 aacaaggttc tggtgaagca aagcgagaac agcaagaagg gcaataggac cccttaccag
     1201 tacctgagca gcaacgagtc caagatctct tacaaccagt tcaagcagca cattctgaac
     1261 ctgtctaagg ccaaagatag aatcagcaag aagaaacgag atatgctgct ggaagaacgg
     1321 gacatcaaca aattcgaggt gcagaaggaa ttcatcaaca gaaaccttgt ggacacccgg
     1381 tacgccactc gggagctgag caacctgctg aagacctact tcagcacaca cgactacgcc
     1441 gtgaaagtga agaccatcaa cggcggcttc acaaaccacc tgaggaaggt gtgggacttc
     1501 aagaagcacc ggaaccacgg ctacaagcac cacgccgagg atgccctggt catcgccaac
     1561 gccgactttc tgttcaaaac ccacaaggcc ctgagacgga cagacaagat cctggaacag
     1621 cctggactgg aagtcaacga caccaccgtg aaggtggaca cagaggagaa gtaccaggag
     1681 ttattcgaga caccgaaaca agtgaagaac atcaagcagt ttagagattt caagtattct
     1741 cacagagttg acaagaagcc caaccggcag ctgatcaatg ataccctgta cagtaccaga
     1801 gagatcgatg gcgaaaccta cgtggtccaa acactgaaag acctgtacgc caaggacaat
     1861 gaaaaggtga aaaagctctt tacagaacgg cctcaaaaga tactgatgta ccagcacgat
     1921 cctaagacct ttgagaaact gatgaccatt ctgaatcagt acgctgaggc aaagaatcct
     1981 ctggccgctt attacgagga taagggcgaa tacgtgacca agtacgccaa gaagggcaac
     2041 ggccctgcca tccacaagat caaatacatc gacaagaaac tgggcagcta cctggacgtg
     2101 agtaacaaat atcctgagac acagaacaag ctggtgaaac tgtctctgaa gagctttaga
     2161 ttcgacatct acaaatgtga acagggctac aagatggtgt ccattggcta cctcgacgta
     2221 ctgaagaagg acaactacta ctacatcccc aaagataagt acgaggccga gaagcagaaa
     2281 aagaagatca aggaaagcga cctcttcgtg ggcagcttct actacaacga cctgatcatg
     2341 tacgaggacg aactcttccg ggtgatcgga gtgaactccg atatcaacaa cctggttgag
     2401 ctgaatatgg tcgacatcac ctacaaggat ttctgcgagg tgaacaacgt gacaggcgag
     2461 aagagaatca agaaaaccat cggaaagaga gtggtgctga tcgagaagta tacgaccgac
     2521 atcctgggaa atctgtataa aacgcccctg cctaagaagc cccagctcat tttcaagaga
     2581 ggcgagctgt ctggcggctc aaaaagaacc gccgacggca gcgaattcga gcccaagaag
     2641 aagaggaaag tctaatagat ctcgactgtg ccttctagtt gccagccatc tgttgtttgc
     2701 ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa
     2761 aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg
     2821 gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggatgcggtg
     2881 ggctctatgg ctcgagcggc ccaagcttaa aaaaatctcg ccaacaagtt gacgagataa
     2941 acacggcatt ttgccttgtt ttagtagatt ctgtaatttt cattacagag tactaaaacc
     3001 aggcatggtt tggtaagtag cggtgtttcg tcctttccac aagatatata aagccaagaa
     3061 atcgaaatac tttcaagtta cggtaagcat atgatagtcc attttaaaac ataattttaa
     3121 aactgcaaac tacccaagaa attattactt tctacgtcac gtattttgta ctaatatctt
     3181 tgtgtttaca gtcaaattaa ttccaattat ctctctaaca gccttgtatc gtatatgcaa
     3241 atatgaagga atcatgggaa ataggccctc ggtaccggcg atgctggctg ccatgaacaa
     3301 aggttggcta taaagaggtc atcagtatat gaaacagccc cctgctgtcc attccttatt
     3361 ccatagaaaa gccttgactt gaggttagat tttttttata ttttgttttg tgttattttt
     3421 ttctttaaca tccctaaaat tttccttaca tgttttacta gccagatttt tcctcctctc
     3481 ctgactactc ccagtcatag ctgtccctct tctcttatgg agatcggatc cgaattcccg
     3541 ataaggatct tcctagagca tggctacgta gataagtagc atggcgggtt aatcattaac
     3601 tacaaggaac ccctagtgat ggagttggcc actccctctc tgcgcgctcg ctcgctcact
     3661 gaggccgggc gaccaaaggt cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc
     3721 gagcgagcgc gcagccttaa ttaacctaat tcactggccg tcgttttaca acgtcgtgac
     3781 tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc
     3841 tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat
     3901 ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     3961 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     4021 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     4081 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     4141 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     4201 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     4261 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     4321 caaaaattta acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc
     4381 ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc
     4441 cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga
     4501 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     4561 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     4621 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     4681 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
     4741 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     4801 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     4861 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     4921 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     4981 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     5041 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     5101 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     5161 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     5221 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     5281 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     5341 tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa
     5401 aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca
     5461 aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag
     5521 gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac
     5581 cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa
     5641 ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc
     5701 accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag
     5761 tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac
     5821 cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc
     5881 gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc
     5941 ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca
     6001 cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc
     6061 tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg
     6121 ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct
     6181 ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata
     6241 ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc
     6301 gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg
     6361 acaggtttcc cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca
     6421 ctcattaggc accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg
     6481 tgagcggata acaatttcac acaggaaaca gctatgacca tgattacgcc agatttaatt
     6541 aagg
//
