LOCUS       sequence-358232-        5779 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV vector carrying a guide RNA targeting the human CTNNB1 mRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5779)
  AUTHORS   Katrekar D, Yen J, Xiang Y, Saha A, Meluzzi D, Savva Y, Mali P
  TITLE     Efficient in vitro and in vivo RNA editing via recruitment of 
            endogenous ADARs using circular guide RNAs.
  JOURNAL   Nat Biotechnol. 2022 Feb 10. pii: 10.1038/s41587-021-01171-4. doi: 
            10.1038/s41587-021-01171-4.
  PUBMED    35145312
REFERENCE   2  (bases 1 to 5779)
  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: "Nat 
            Biotechnol. 2022 Feb 10. pii: 10.1038/s41587-021-01171-4. doi: 
            10.1038/s41587-021-01171-4."
COMMENT     Sequence Label: Circular 200,100 with interspersed loops (CTNNB1)
FEATURES             Location/Qualifiers
     source          1..5779
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     promoter        160..408
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     160..180
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     331..350
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     promoter        817..1130
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for mouse U6 snRNA (Das 
                     et al., 1988)"
     primer_bind     1049..1069
                     /label=mU6-F
                     /note="Mouse U6 promoter, forward primer"
     protein_bind    complement(1142..1166)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     enhancer        1181..1560
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1561..1764
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1715..1735
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1761..1785
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      1876..1885
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1882..1884,1885..1887,1888..2598)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1882..1884,1885..1887,1888..2598)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1927..1948)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2188..2207)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2535..2556
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(2677..2696)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    2721..2769
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     repeat_region   2984..3124
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2984..3113
                     /label=AAV2 ITR
     primer_bind     complement(3164..3180)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3164..3180)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3177..3199)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3188..3204
                     /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(3212..3218,3219..3236,3237..3242))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3257..3278
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(3566..4154)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3646..3665)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4325..5140)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(4325..5140)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     5048..5067
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(5141..5245)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5313..5331
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5369..5391)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5491..5510
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5704..5726
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5719..5735
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5760..5776
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtggagg agggcctatt tcccatgatt
      181 ccttcatatt tgcatatacg atacaaggct gttagagaga taattagaat taatttgact
      241 gtaaacacaa agatattagt acaaaatacg tgacgtagaa agtaataatt tcttgggtag
      301 tttgcagttt taaaattatg ttttaaaatg gactatcata tgcttaccgt aacttgaaag
      361 tatttcgatt tcttggcttt atatatcttg tggaaaggac gaaacaccgc catcagtcgc
      421 cggtcccaag cccggataaa atgggagggg gcgggaaacc gcctaaccat gccgactgat
      481 ggcagacaaa aaaaaagttt acttcaattg ttcccatagg tttgtcagct tggttagtga
      541 cagaggcact ttctgagaat ggagcccacc cctcgagccc tctcagcaac tccacagggg
      601 tttcacaatg caagttctct gaatacagct gtataatctc agaaaggctg ctattataca
      661 ttaaaacagc aaatgttact catttgagta ttaaaaaaaa aaactgccat cagtcggcgt
      721 ggactgtaga acactgccaa tgccggtccc aagcccggat aaaagtggag ggtacagtcc
      781 acgctttttt cgtactgagt cgcccagtct cagatagatc cgacgccgcc atctctaggc
      841 ccgcgccggc cccctcgcac agacttgtgg gagaagctcg gctactcccc tgccccggtt
      901 aatttgcata taatatttcc tagtaactat agaggcttaa tgtgcgataa aagacagata
      961 atctgttctt tttaatacta gctacatttt acatgatagg cttggatttc tataagagat
     1021 acaaatacta aattattatt ttaaaaaaca gcacaaaagg aaactcaccc taactgtaaa
     1081 gtaattgtgt gttttgagac tataaatatc ccttggagaa aagccttgtt tggattccta
     1141 gacccagctt tcttgtacaa agttggcatt aatacgcgtt gacattgatt attgactagt
     1201 tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt
     1261 acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg
     1321 tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg
     1381 gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt
     1441 acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg
     1501 accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg
     1561 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
     1621 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
     1681 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
     1741 tgggaggtct atataagcag agctcgttta gtgaaccgtc agatcgcctg gagacgccat
     1801 ccacgctgtt ttgacctcca tagaagacac cgggaccgat ccagcctccg gactctagag
     1861 gatcgaaccc ttaaggccac catggtgagc aagggcgagg agctgttcac cggggtggtg
     1921 cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag
     1981 ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag
     2041 ctgcccgtgc cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc
     2101 cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac
     2161 gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg
     2221 aagttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag
     2281 gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc
     2341 atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag
     2401 gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc
     2461 gtgctgctgc ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac
     2521 gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc
     2581 atggacgagc tgtacaagta ctcagatctc gagctcaagt agcttaaggg ttcgatccct
     2641 actggttagt aatgagttta aacgggggag gctaactgaa acacggaagg agacaatacc
     2701 ggaaggaacc cgcgctatga cggcaataaa aagacagaat aaaacgcacg ggtgttgggt
     2761 cgtttgttca taaacgcggg gttcggtccc agggctggca ctctgtcgat accccaccga
     2821 gaccccattg gggccaatac gcccgcgttt cttccttttc cccaccccac cccccaagtt
     2881 cgggtgaagg cccagggctc gcagccaacg tcggggcggc aggccctgcc atagcagatc
     2941 tgcgctgatt ttgtaggtaa ccacgtgcgg accgagcggc cgcaggaacc cctagtgatg
     3001 gagttggcca ctccctctct gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc
     3061 gcccgacgcc cgggctttgc ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg
     3121 caggcttgga tcccaatggc gcgccgagct tggctcgagc atggtcatag ctgtttcctg
     3181 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     3241 aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     3301 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     3361 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     3421 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     3481 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     3541 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     3601 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     3661 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     3721 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     3781 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     3841 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     3901 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     3961 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     4021 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     4081 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     4141 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     4201 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     4261 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     4321 acagttagaa aaactcatcg agcatcaaat gaaactgcaa tttattcata tcaggattat
     4381 caataccata tttttgaaaa agccgtttct gtaatgaagg agaaaactca ccgaggcagt
     4441 tccataggat ggcaagatcc tggtatcggt ctgcgattcc gactcgtcca acatcaatac
     4501 aacctattaa tttcccctcg tcaaaaataa ggttatcaag tgagaaatca ccatgagtga
     4561 cgactgaatc cggtgagaat ggcaaaagtt tatgcatttc tttccagact tgttcaacag
     4621 gccagccatt acgctcgtca tcaaaatcac tcgcatcaac caaaccgtta ttcattcgtg
     4681 attgcgcctg agcgagacga aatacgcgat cgctgttaaa aggacaatta caaacaggaa
     4741 tcgaatgcaa ccggcgcagg aacactgcca gcgcatcaac aatattttca cctgaatcag
     4801 gatattcttc taatacctgg aatgctgttt tcccagggat cgcagtggtg agtaaccatg
     4861 catcatcagg agtacggata aaatgcttga tggtcggaag aggcataaat tccgtcagcc
     4921 agtttagtct gaccatctca tctgtaacat cattggcaac gctacctttg ccatgtttca
     4981 gaaacaactc tggcgcatcg ggcttcccat acaatcgata gattgtcgca cctgattgcc
     5041 cgacattatc gcgagcccat ttatacccat ataaatcagc atccatgttg gaatttaatc
     5101 gcggcctaga gcaagacgtt tcccgttgaa tatggctcat actcttcctt tttcaatatt
     5161 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     5221 aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtctaag
     5281 aaaccattat tatcatgaca ttaacctata aaaataggcg tatcacgagg ccctttcgtc
     5341 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
     5401 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
     5461 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
     5521 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
     5581 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
     5641 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
     5701 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt gacgcgccat tgggatgttg
     5761 taaaacgacg gccagtgaa
//
