LOCUS       sequence-375708-        6345 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Deliver a split compact Prime Editor in vivo.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6345)
  AUTHORS   Zheng C, Liang SQ, Liu B, Liu P, Kwan SY, Wolfe SA, Xue W
  TITLE     A flexible split prime editor using truncated reverse transcriptase 
            improves dual-AAV delivery in mouse liver.
  JOURNAL   Mol Ther. 2022 Mar 2;30(3):1343-1351. doi: 
            10.1016/j.ymthe.2022.01.005. Epub 2022 Jan 5.
  PUBMED    34998953
REFERENCE   2  (bases 1 to 6345)
  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"; doi: 
            "10.1016/j.ymthe.2022.01.005"; journalName: "Mol Ther"; date: 
            "2022-03-2- 2"; volume: "30"; issue: "3"; pages: "1343-1351"
COMMENT     Sequence Label: AAV8-cPE-N
FEATURES             Location/Qualifiers
     source          1..6345
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        177..425
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        447..522
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        612..860
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        882..957
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     regulatory      1295..1304
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1310..1336
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             1337..3052
                     /codon_start=1
                     /product="N-terminal portion of Streptococcus pyogenes Cas9
                     (Zetsche et al., 2015)"
                     /label=Cas9(N)
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIE"
     primer_bind     complement(3386..3403)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3392..3599
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3608..3748
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3608..3737
                     /label=AAV2 ITR
     rep_origin      3823..4278
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3910..3929)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4120..4141
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4295..4314)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4414..4436
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4474..4492)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4560..4664
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4665..4733,4734..5525)
                     /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(4883..4902)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5696..6284
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6185..6204
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggcctcta gaagagtaat tcaaccccaa acaacaacgt ttaaacgagg
      181 gcctatttcc catgattcct tcatatttgc atatacgata caaggctgtt agagagataa
      241 ttggaattaa tttgactgta aacacaaaga tattagtaca aaatacgtga cgtagaaagt
      301 aataatttct tgggtagttt gcagttttaa aattatgttt taaaatggac tatcatatgc
      361 ttaccgtaac ttgaaagtat ttcgatttct tggctttata tatcttgtgg aaaggacgaa
      421 acaccgttgc tgctactgtg ccccacgttt tagagctaga aatagcaagt taaaataagg
      481 ctagtccgtt atcaacttga aaaagtggga ccgagtcggt ccagcgccgg ttcaggggca
      541 cagtagcatt tttttgaatt cgatatcaag cttatcgata ccgtcgagct ggagctccac
      601 cccccttcac cgagggccta tttcccatga ttccttcata tttgcatata cgatacaagg
      661 ctgttagaga gataattgga attaatttga ctgtaaacac aaagatatta gtacaaaata
      721 cgtgacgtag aaagtaataa tttcttgggt agtttgcagt tttaaaatta tgttttaaaa
      781 tggactatca tatgcttacc gtaacttgaa agtatttcga tttcttggct ttatatatct
      841 tgtggaaagg acgaaacacc ggccatcctc ctgggacggg agttttagag ctagaaatag
      901 caagttaaaa taaggctagt ccgttatcaa cttgaaaaag tggcaccgag tcggtgcttt
      961 ttttgaattc gatatcaagc ttatcgatac cgtcgggtac ctctagaccg cggcgcgcct
     1021 ccatggatat caagcttatg gaggcggtac tatgtagatg agaattcagg agcaaactgg
     1081 gaaaagcaac tgcttccaaa tatttgtgat ttttacagtg tagttttgga aaaactctta
     1141 gcctaccaat tcttctaagt gttttaaaat gtgggagcca gtacacatga agttatagag
     1201 tgttttaatg aggcttaaat atttaccgta actatgaaat gctacgcata tcatgctgtt
     1261 caggctccgt ggccacgcaa ctcatactac cggtgccacc atggctagcc ccgccgccaa
     1321 gcgcgtgaag ctggacgaca agaagtacag catcggcctg gacatcggca ccaactctgt
     1381 gggctgggcc gtgatcaccg acgagtacaa ggtgcccagc aagaaattca aggtgctggg
     1441 caacaccgac cggcacagca tcaagaagaa cctgatcgga gccctgctgt tcgacagcgg
     1501 cgaaacagcc gaggccaccc ggctgaagag aaccgccaga agaagataca ccagacggaa
     1561 gaaccggatc tgctatctgc aagagatctt cagcaacgag atggccaagg tggacgacag
     1621 cttcttccac agactggaag agtccttcct ggtggaagag gataagaagc acgagcggca
     1681 ccccatcttc ggcaacatcg tggacgaggt ggcctaccac gagaagtacc ccaccatcta
     1741 ccacctgaga aagaaactgg tggacagcac cgacaaggcc gacctgcggc tgatctatct
     1801 ggccctggcc cacatgatca agttccgggg ccacttcctg atcgagggcg acctgaaccc
     1861 cgacaacagc gacgtggaca agctgttcat ccagctggtg cagacctaca accagctgtt
     1921 cgaggaaaac cccatcaacg ccagcggcgt ggacgccaag gccatcctgt ctgccagact
     1981 gagcaagagc agacggctgg aaaatctgat cgcccagctg cccggcgaga agaagaatgg
     2041 cctgttcgga aacctgattg ccctgagcct gggcctgacc cccaacttca agagcaactt
     2101 cgacctggcc gaggatgcca aactgcagct gagcaaggac acctacgacg acgacctgga
     2161 caacctgctg gcccagatcg gcgaccagta cgccgacctg tttctggccg ccaagaacct
     2221 gtccgacgcc atcctgctga gcgacatcct gagagtgaac accgagatca ccaaggcccc
     2281 cctgagcgcc tctatgatca agagatacga cgagcaccac caggacctga ccctgctgaa
     2341 agctctcgtg cggcagcagc tgcctgagaa gtacaaagag attttcttcg accagagcaa
     2401 gaacggctac gccggctaca ttgacggcgg agccagccag gaagagttct acaagttcat
     2461 caagcccatc ctggaaaaga tggacggcac cgaggaactg ctcgtgaagc tgaacagaga
     2521 ggacctgctg cggaagcagc ggaccttcga caacggcagc atcccccacc agatccacct
     2581 gggagagctg cacgccattc tgcggcggca ggaagatttt tacccattcc tgaaggacaa
     2641 ccgggaaaag atcgagaaga tcctgacctt ccgcatcccc tactacgtgg gccctctggc
     2701 caggggaaac agcagattcg cctggatgac cagaaagagc gaggaaacca tcaccccctg
     2761 gaacttcgag gaagtggtgg acaagggcgc ttccgcccag agcttcatcg agcggatgac
     2821 caacttcgat aagaacctgc ccaacgagaa ggtgctgccc aagcacagcc tgctgtacga
     2881 gtacttcacc gtgtataacg agctgaccaa agtgaaatac gtgaccgagg gaatgagaaa
     2941 gcccgccttc ctgagcggcg agcagaaaaa ggccatcgtg gacctgctgt tcaagaccaa
     3001 ccggaaagtg accgtgaagc agctgaaaga ggactacttc aagaaaatcg agtgcctgtc
     3061 ctacgagaca gagatcctga cagtggagta tggcctgctg ccaatcggca agatcgtgga
     3121 gaagaggatc gagtgtaccg tgtactctgt ggataacaat ggcaacatct atacacagcc
     3181 cgtggcacag tggcacgata ggggagagca ggaggtgttc gagtattgcc tggaggacgg
     3241 cagcctgatc agggcaacca aggaccacaa gttcatgaca gtggatggcc agatgctgcc
     3301 catcgacgag attttcgagc gggagctgga cctgatgaga gtggataacc tgcctaattg
     3361 agaattccta gagctcgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt
     3421 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     3481 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt
     3541 ggggtggggc aggacagcaa gggggaggat tgggaagaga atagcaggca tgctggggag
     3601 cggccgcagg aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc
     3661 actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg
     3721 agcgagcgag cgcgcagctg cctgcagggg cgcctgatgc ggtattttct ccttacgcat
     3781 ctgtgcggta tttcacaccg catacgtcaa agcaaccata gtacgcgccc tgtagcggcg
     3841 cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgcct
     3901 tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc
     3961 gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg
     4021 accccaaaaa acttgatttg ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg
     4081 tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg
     4141 gaacaacact caactctatc tcgggctatt cttttgattt ataagggatt ttgccgattt
     4201 cggtctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa
     4261 tattaacgtt tacaatttta tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     4321 taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc
     4381 cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt
     4441 caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg
     4501 ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc
     4561 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     4621 aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt
     4681 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     4741 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     4801 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     4861 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc
     4921 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     4981 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     5041 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     5101 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     5161 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa
     5221 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     5281 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     5341 gctggtttat tgctgataaa tctggagccg gtgagcgtgg aagccgcggt atcattgcag
     5401 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     5461 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     5521 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt
     5581 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     5641 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     5701 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     5761 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     5821 gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga
     5881 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     5941 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     6001 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     6061 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     6121 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     6181 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     6241 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     6301 cctttttacg gttcctggcc ttttgctggc cttttgctca catgt
//
