LOCUS       sequence-488810-        9699 bp    DNA     circular SYN 08-OCT-2025
DEFINITION  Encodes second generation porcine endogenous retrovirus-derived 
            prime editing (pvPE) system into mammalian cells for gene editing..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9699)
  AUTHORS   Liu W, Duan W, Peng Z, Liao Y, Wang X, Liu R, Jing Q, Jiang H, Fan 
            Y, Ge L, Huang L, Xing Y
  TITLE     Highly efficient prime editors for mammalian genome editing based on
            porcine retrovirus reverse transcriptase.
  JOURNAL   Trends Biotechnol. 2025 Aug 30:S0167-7799(25)00314-2. doi: 
            10.1016/j.tibtech.2025.07.029.
  PUBMED    40885667
REFERENCE   2  (bases 1 to 9699)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Oct 8, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Trends 
            Biotechnol. 2025 Aug 30:S0167-7799(25)00314-2. doi: 
            10.1016/j.tibtech.2025.07.029."
COMMENT     Sequence Label: pvPE-V2
FEATURES             Location/Qualifiers
     source          1..9699
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             694..714
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(715..3228,3229..3231,3232..4815)
                     /codon_start=1
                     /product="nickase mutant of the Cas9 endonuclease from the 
                     Streptococcus pyogenes Type II CRISPR/Cas system"
                     /label=Cas9(H840A)
                     /note="generates RNA-guided single strand nicks in DNA due 
                     to the H840A mutation in the HNH catalytic domain"
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             715..4815
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             6934..6954
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6963..6980
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(7003..7020)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    7009..7233
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(7304..7320)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7304..7320)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7317..7339)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7328..7344
                     /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(7352..7358,7359..7376,7377..7382))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7397..7418
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(7535..7552)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7706..8294)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7786..7805)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(8465..9256,9257..9325))
                     /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     9088..9107
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9326..9430)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(9509..9528)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc
      601 agatccgcta gagatccgcg gccgctaata cgactcacta tagggagagc cgccaccatg
      661 aaacggacag ccgacggaag cgagttcgag tcaccaaaga agaagcggaa agtcgacaag
      721 aagtacagca tcggcctgga catcggcacc aactctgtgg gctgggccgt gatcaccgac
      781 gagtacaagg tgcccagcaa gaaattcaag gtgctgggca acaccgaccg gcacagcatc
      841 aagaagaacc tgatcggagc cctgctgttc gacagcggcg aaacagccga ggccacccgg
      901 ctgaagagaa ccgccagaag aagatacacc agacggaaga accggatctg ctatctgcaa
      961 gagatcttca gcaacgagat ggccaaggtg gacgacagct tcttccacag actggaagag
     1021 tccttcctgg tggaagagga taagaagcac gagcggcacc ccatcttcgg caacatcgtg
     1081 gacgaggtgg cctaccacga gaagtacccc accatctacc acctgagaaa gaaactggtg
     1141 gacagcaccg acaaggccga cctgcggctg atctatctgg ccctggccca catgatcaag
     1201 ttccggggcc acttcctgat cgagggcgac ctgaaccccg acaacagcga cgtggacaag
     1261 ctgttcatcc agctggtgca gacctacaac cagctgttcg aggaaaaccc catcaacgcc
     1321 agcggcgtgg acgccaaggc catcctgtct gccagactga gcaagagcag acggctggaa
     1381 aatctgatcg cccagctgcc cggcgagaag aagaatggcc tgttcggaaa cctgattgcc
     1441 ctgagcctgg gcctgacccc caacttcaag agcaacttcg acctggccga ggatgccaaa
     1501 ctgcagctga gcaaggacac ctacgacgac gacctggaca acctgctggc ccagatcggc
     1561 gaccagtacg ccgacctgtt tctggccgcc aagaacctgt ccgacgccat cctgctgagc
     1621 gacatcctga gagtgaacac cgagatcacc aaggcccccc tgagcgcctc tatgatcaag
     1681 agatacgacg agcaccacca ggacctgacc ctgctgaaag ctctcgtgcg gcagcagctg
     1741 cctgagaagt acaaagagat tttcttcgac cagagcaaga acggctacgc cggctacatt
     1801 gacggcggag ccagccagga agagttctac aagttcatca agcccatcct ggaaaagatg
     1861 gacggcaccg aggaactgct cgtgaagctg aacagagagg acctgctgcg gaagcagcgg
     1921 accttcgaca acggcagcat cccccaccag atccacctgg gagagctgca cgccattctg
     1981 cggcggcagg aagattttta cccattcctg aaggacaacc gggaaaagat cgagaagatc
     2041 ctgaccttcc gcatccccta ctacgtgggc cctctggcca ggggaaacag cagattcgcc
     2101 tggatgacca gaaagagcga ggaaaccatc accccctgga acttcgagga agtggtggac
     2161 aagggcgctt ccgcccagag cttcatcgag cggatgacca acttcgataa gaacctgccc
     2221 aacgagaagg tgctgcccaa gcacagcctg ctgtacgagt acttcaccgt gtataacgag
     2281 ctgaccaaag tgaaatacgt gaccgaggga atgagaaagc ccgccttcct gagcggcgag
     2341 cagaaaaagg ccatcgtgga cctgctgttc aagaccaacc ggaaagtgac cgtgaagcag
     2401 ctgaaagagg actacttcaa gaaaatcgag tgcttcgact ccgtggaaat ctccggcgtg
     2461 gaagatcggt tcaacgcctc cctgggcaca taccacgatc tgctgaaaat tatcaaggac
     2521 aaggacttcc tggacaatga ggaaaacgag gacattctgg aagatatcgt gctgaccctg
     2581 acactgtttg aggacagaga gatgatcgag gaacggctga aaacctatgc ccacctgttc
     2641 gacgacaaag tgatgaagca gctgaagcgg cggagataca ccggctgggg caggctgagc
     2701 cggaagctga tcaacggcat ccgggacaag cagtccggca agacaatcct ggatttcctg
     2761 aagtccgacg gcttcgccaa cagaaacttc atgcagctga tccacgacga cagcctgacc
     2821 tttaaagagg acatccagaa agcccaggtg tccggccagg gcgatagcct gcacgagcac
     2881 attgccaatc tggccggcag ccccgccatt aagaagggca tcctgcagac agtgaaggtg
     2941 gtggacgagc tcgtgaaagt gatgggccgg cacaagcccg agaacatcgt gatcgaaatg
     3001 gccagagaga accagaccac ccagaaggga cagaagaaca gccgcgagag aatgaagcgg
     3061 atcgaagagg gcatcaaaga gctgggcagc cagatcctga aagaacaccc cgtggaaaac
     3121 acccagctgc agaacgagaa gctgtacctg tactacctgc agaatgggcg ggatatgtac
     3181 gtggaccagg aactggacat caaccggctg tccgactacg atgtggacgc tatcgtgcct
     3241 cagagctttc tgaaggacga ctccatcgac aacaaggtgc tgaccagaag cgacaagaac
     3301 cggggcaaga gcgacaacgt gccctccgaa gaggtcgtga agaagatgaa gaactactgg
     3361 cggcagctgc tgaacgccaa gctgattacc cagagaaagt tcgacaatct gaccaaggcc
     3421 gagagaggcg gcctgagcga actggataag gccggcttca tcaagagaca gctggtggaa
     3481 acccggcaga tcacaaagca cgtggcacag atcctggact cccggatgaa cactaagtac
     3541 gacgagaatg acaagctgat ccgggaagtg aaagtgatca ccctgaagtc caagctggtg
     3601 tccgatttcc ggaaggattt ccagttttac aaagtgcgcg agatcaacaa ctaccaccac
     3661 gcccacgacg cctacctgaa cgccgtcgtg ggaaccgccc tgatcaaaaa gtaccctaag
     3721 ctggaaagcg agttcgtgta cggcgactac aaggtgtacg acgtgcggaa gatgatcgcc
     3781 aagagcgagc aggaaatcgg caaggctacc gccaagtact tcttctacag caacatcatg
     3841 aactttttca agaccgagat taccctggcc aacggcgaga tccggaagcg gcctctgatc
     3901 gagacaaacg gcgaaaccgg ggagatcgtg tgggataagg gccgggattt tgccaccgtg
     3961 cggaaagtgc tgagcatgcc ccaagtgaat atcgtgaaaa agaccgaggt gcagacaggc
     4021 ggcttcagca aagagtctat cctgcccaag aggaacagcg ataagctgat cgccagaaag
     4081 aaggactggg accctaagaa gtacggcggc ttcgacagcc ccaccgtggc ctattctgtg
     4141 ctggtggtgg ccaaagtgga aaagggcaag tccaagaaac tgaagagtgt gaaagagctg
     4201 ctggggatca ccatcatgga aagaagcagc ttcgagaaga atcccatcga ctttctggaa
     4261 gccaagggct acaaagaagt gaaaaaggac ctgatcatca agctgcctaa gtactccctg
     4321 ttcgagctgg aaaacggccg gaagagaatg ctggcctctg ccggcgaact gcagaaggga
     4381 aacgaactgg ccctgccctc caaatatgtg aacttcctgt acctggccag ccactatgag
     4441 aagctgaagg gctcccccga ggataatgag cagaaacagc tgtttgtgga acagcacaag
     4501 cactacctgg acgagatcat cgagcagatc agcgagttct ccaagagagt gatcctggcc
     4561 gacgctaatc tggacaaagt gctgtccgcc tacaacaagc accgggataa gcccatcaga
     4621 gagcaggccg agaatatcat ccacctgttt accctgacca atctgggagc ccctgccgcc
     4681 ttcaagtact ttgacaccac catcgaccgg aagaggtaca ccagcaccaa agaggtgctg
     4741 gacgccaccc tgatccacca gagcatcacc ggcctgtacg agacacggat cgacctgtct
     4801 cagctgggag gtgactctgg aggatctagc ggaggatcct ctggcagcga gacaccagga
     4861 acaagcgagt cagcaacacc agagagcagt ggcggcagca gcggcggcag cagcaccctg
     4921 cagctggatg atgaatacag actatacagt cctctggtga agccagacca gaatatccag
     4981 ttctggctcg agcagttccc tcaggcctgg gctgaaaccg ccggagtggg cctggccaaa
     5041 caggtgcccc ctcaagtcat tcagctcaag gcctctgcca ctccagtgtc tgtgcggcag
     5101 agaccgctgt ctaaggaagc tagagaaggc atccggccac acgtacaaag actgatacag
     5161 cagggcatcc tggtgcctgt tcagagcccc tggaacaccc ccctgctgcc tgtgagaaaa
     5221 cctggaacaa atgattaccg gcctgtgcag gacctgagag aggtcaacaa acgggtgcag
     5281 gatatccatc ctaccgtgcc caacccctat aacctgctgt gtgccctgcc accacagcgg
     5341 agctggtaca cagttctgga tctgaaggat gccttcttct gtctgcggct ccacccaacc
     5401 agccagcctc tctttgcttt tgaatggcgg gaccctggcg ccggccgaac gggccagcta
     5461 acctggacaa gactgcccca gggttttaaa aactctccca ccatctttaa tgaagccctt
     5521 cacagggacc tcgccaattt tagaatccag caccctcaag taactctgct ccagtatgct
     5581 gatgacctgc tacttgccgg cgccaccaag caagactgtt tggaaggaac caaagccctg
     5641 ctgctggagc tgtctgacct aggctaccga gcaagtgcta agaaggcaca gatctgccgg
     5701 cgagaggtca catacctagg atacagcctc agaggcggcc agaggtggct caccgaggcc
     5761 aggaagagga ccgtggtgca gatcccagcc ccaaccacgg caaagcaggt gagggaattt
     5821 ctggggaagg ccgggttctg cagactgttc attcctggct tcgccacctt ggcggctccc
     5881 ctctacccac ttacaaagga gaagggcgag ttctcctggg cgccagagca tcagaaggcc
     5941 tttgacgcca tcaaaaaggc tctgctgtct gccccagccc tggccttacc agatgtcacc
     6001 aagccgttca ccctgtatgt tgatgaaaga aaaggggtcg cgcggggcgt actgacccag
     6061 actctgggcc catggagacg gcctgtggcc tacctaagca agaagctgga ccctgttgcc
     6121 tcaggctggc ccgtgcgtct gaaagccata gctgctgtgg ccatcctggt gaaggatgca
     6181 gacaagctaa ctctaggtca gaacatcacc gtcatcgccc cccacgctct ggaaaacatc
     6241 gtgcgccagc ccccagaccg ctggatgacc aacgcccgca tgacacacta tcagagcctc
     6301 ttactgacag agagggtgac gtttgctcct cccgccgccc tcaatcctgc caccctgcta
     6361 cctgaagaaa ccgacgaacc cgtgacccac gactgccacc agttgctcat cgaggaaaca
     6421 ggagtcagga aagacttgac ggacatccct ctaactgggg aggttctgac atggttcaca
     6481 gatgggagca gctacgtggt ggagggaaag aggatggcgg gcgcagccgt ggtggacggc
     6541 acccgtacta tttgggctag ctccttacct gagggtacat cggcgcagaa ggccgagctt
     6601 atggccttaa cacaagctct gcggctggcg gaaggcaaga gcataaacat ctacacagat
     6661 agccgctacg ccttcgccac cgcgcatgtc cacggagcca tctataaaca gagaggttgg
     6721 ctgacctcag caggccgaga aatcaagaat aaggaggaga tcctcagcct gctggaggct
     6781 ctgcacctgc ccaaaagact ggccatcatc cactgcccgg gccaccaaaa ggccaaggac
     6841 cccatctcca gaggtaacca gatggctgac cgagtggcaa agcaggctgc ctctggcggc
     6901 tcaaaaagaa ccgccgacgg cagcgaattc gagcccaaga agaagaggaa agtctaaccg
     6961 gtcatcatca ccatcaccat tgagtttaaa cccgctgatc agcctcgact gtgccttcta
     7021 gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca
     7081 ctcccactgt cctttcctaa taaaatgaga aaattgcatc gcattgtctg agtaggtgtc
     7141 attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata
     7201 gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg
     7261 gctcgatacc gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg
     7321 tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta
     7381 aagcctaggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg
     7441 ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga
     7501 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     7561 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     7621 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     7681 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     7741 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     7801 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     7861 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     7921 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     7981 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     8041 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     8101 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     8161 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     8221 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     8281 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacact cagtggaacg
     8341 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     8401 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     8461 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     8521 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     8581 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     8641 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     8701 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     8761 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     8821 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     8881 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     8941 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     9001 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     9061 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     9121 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     9181 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     9241 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     9301 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     9361 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     9421 gggttccgcg cacatttccc cgaaaagtgc cacctgacgt cgacggatcg ggagatcgat
     9481 ctcccgatcc cctagggtct actctcagta caatctgctc tgatgccgca tagttaagcc
     9541 agtatctgct ccctgcttgt gtgttggagg tcgctgagta gtgcgcgagc aaaatttaag
     9601 ctacaacaag gcaaggcttg accgacaatt gcatgaagaa tctgcttagg gttaggcgtt
     9661 ttgcgctgct tcgcgatgta cgggccagat atacgcgtt
//
