LOCUS       sequence-350237-       14038 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  piggyBac transposon containing both PE2 and a pegRNA cloning site..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14038)
  AUTHORS   Wolff JH, Haldrup J, Thomsen EA, Andersen S, Mikkelsen JG
  TITLE     piggyPrime: High-Efficacy Prime Editing in Human Cells Using 
            piggyBac-Based DNA Transposition.
  JOURNAL   Front Genome Ed. 2021 Nov 12;3:786893. doi: 
            10.3389/fgeed.2021.786893. eCollection 2021.
  PUBMED    34870275
REFERENCE   2  (bases 1 to 14038)
  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: "Front 
            Genome Ed."; date: "2021-11-12"; volume: "3:786893. doi"; pages: " 
            10.3389/fgeed.2021.786893. eCollection 2021"
COMMENT     Sequence Label: pPBT-PE2-PuroTK-pegRNA_GG
FEATURES             Location/Qualifiers
     source          1..14038
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    88..109
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(124..129,130..147,148..154)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    162..178
                     /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     167..189
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     186..202
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     186..202
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     repeat_region   693..727
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     enhancer        1008..1387
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1388..1591
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1542..1562
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     1633..1652
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1633..1651
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             1701..1721
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(1722..4235,4236..4238,4239..5822)
                     /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             1722..5822
                     /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             7995..8015
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8025..8081
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             8082..8675
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
                     TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
                     QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
                     PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     8575..8595
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             8682..9677
                     /codon_start=1
                     /product="truncated herpes simplex virus thymidine kinase 
                     (Salomon et al., 1995)"
                     /label=Delta-TK
                     /note="confers sensitivity to nucleoside analogs such as 
                     acyclovir or ganciclovir"
                     /translation="MPTLLRVYIDGPHGMGKTTTTQLLVALGSRDDIVYVPEPMTYWQV
                     LGASETIANIYTTQHRLDQGEISAGDAAVVMTSAQITMGMPYAVTDAVLAPHIGGEAGS
                     SHAPPPALTLIFDRHPIAALLCYPAARYLMGSMTPQAVLAFVALIPPTLPGTNIVLGAL
                     PEDRHIDRLAKRQRPGERLDLAMLAAIRRVYGLLANTVRYLQGGGSWREDWGQLSGTAV
                     PPQGAEPQSNAGPRPHIGDTLFTLFRAPELLAPNGDLYNVFAWALDVLAKRLRPMHVFI
                     LDYDQSPAGCRDALLQLTSGMVQTHVTTPGSIPTICDLARTFAREMGEAN"
     primer_bind     complement(9704..9721)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9710..9934
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     9943..9959
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     9943..9959
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     promoter        10015..10263
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     10015..10035
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     10186..10205
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     terminator      complement(10273..10304)
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     CDS             complement(10321..10998)
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     RBS             11005..11016
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     protein_bind    11034..11052
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    11059..11077
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     repeat_region   11283..11345
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(11475..11492)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(11475..11491)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11484..11506)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      11633..12088
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(11720..11739)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     11930..11951
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        12114..12218
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(12219..12287,12288..13079)
                     /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(12437..12456)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      13250..13838
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     13739..13758
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     13992..14009
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt ttcccgactg
       61 gaaagcgggc agtgagcgca acgcaattaa tgtgagttag ctcactcatt aggcacccca
      121 ggctttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt
      181 tcacacagga aacagctatg accatgatta cgccaagcgc gcccgccggg taactcacgg
      241 ggtatccatg tccatttctg cggcatccag ccaggatacc cgtcctcgct gacgtaatat
      301 cccagcgccg caccgctgtc attaatctgc acaccggcac ggcagttccg gctgtcgccg
      361 gtattgttcg ggttgctgat gcgcttcggg ctgaccatcc ggaactgtgt ccggaaaagc
      421 cgcgacgaac tggtatccca ggtggcctga acgaacagtt caccgttaaa ggcgtgcatg
      481 gccacacctt cccgaatcat catggtaaac gtgcgttttc gctcaacgtc aatgcagcag
      541 cagtcatcct cggcaaactc tttccatgcc gcttcaacct cgcgggaaaa ggcacgggct
      601 tcttcctccc cgatgcccag atagcgccag cttgggcgat gactgagccg gaaaaaagac
      661 ccgacgatat gatcctgatg cagctagatt aaccctagaa agatagtctg cgtaaaattg
      721 acgcatgcat tcttgaaata ttgctctctc tttctaaata gcgcgaatcc gtcgctgtgc
      781 atttaggaca tctcagtcgc cgcttggagc tcccgtgagg cgtgcttgtc aatgcggtaa
      841 gtgtcactga ttttgaacta taacgaccgc gtgagtcaaa atgacgcatg attatctttt
      901 acgtgacttt taagatttaa ctcatacgat aattatattg ttatttcatg ttctacttac
      961 gtgataactt attatatata tattttcttg ttatagatat caagcttgac attgattatt
     1021 gactagttat taatagtaat caattacggg gtcattagtt catagcccat atatggagtt
     1081 ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc
     1141 attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg
     1201 tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat
     1261 gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca
     1321 gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat
     1381 taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt ttgactcacg
     1441 gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc accaaaatca
     1501 acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg gcggtaggcg
     1561 tgtacggtgg gaggtctata taagcagagc tggtttagtg aaccgtcaga tccgctagag
     1621 atccgcggcc gctaatacga ctcactatag ggagagccgc caccatgaaa cggacagccg
     1681 acggaagcga gttcgagtca ccaaagaaga agcggaaagt cgacaagaag tacagcatcg
     1741 gcctggacat cggcaccaac tctgtgggct gggccgtgat caccgacgag tacaaggtgc
     1801 ccagcaagaa attcaaggtg ctgggcaaca ccgaccggca cagcatcaag aagaacctga
     1861 tcggagccct gctgttcgac agcggcgaaa cagccgaggc cacccggctg aagagaaccg
     1921 ccagaagaag atacaccaga cggaagaacc ggatctgcta tctgcaagag atcttcagca
     1981 acgagatggc caaggtggac gacagcttct tccacagact ggaagagtcc ttcctggtgg
     2041 aagaggataa gaagcacgag cggcacccca tcttcggcaa catcgtggac gaggtggcct
     2101 accacgagaa gtaccccacc atctaccacc tgagaaagaa actggtggac agcaccgaca
     2161 aggccgacct gcggctgatc tatctggccc tggcccacat gatcaagttc cggggccact
     2221 tcctgatcga gggcgacctg aaccccgaca acagcgacgt ggacaagctg ttcatccagc
     2281 tggtgcagac ctacaaccag ctgttcgagg aaaaccccat caacgccagc ggcgtggacg
     2341 ccaaggccat cctgtctgcc agactgagca agagcagacg gctggaaaat ctgatcgccc
     2401 agctgcccgg cgagaagaag aatggcctgt tcggaaacct gattgccctg agcctgggcc
     2461 tgacccccaa cttcaagagc aacttcgacc tggccgagga tgccaaactg cagctgagca
     2521 aggacaccta cgacgacgac ctggacaacc tgctggccca gatcggcgac cagtacgccg
     2581 acctgtttct ggccgccaag aacctgtccg acgccatcct gctgagcgac atcctgagag
     2641 tgaacaccga gatcaccaag gcccccctga gcgcctctat gatcaagaga tacgacgagc
     2701 accaccagga cctgaccctg ctgaaagctc tcgtgcggca gcagctgcct gagaagtaca
     2761 aagagatttt cttcgaccag agcaagaacg gctacgccgg ctacattgac ggcggagcca
     2821 gccaggaaga gttctacaag ttcatcaagc ccatcctgga aaagatggac ggcaccgagg
     2881 aactgctcgt gaagctgaac agagaggacc tgctgcggaa gcagcggacc ttcgacaacg
     2941 gcagcatccc ccaccagatc cacctgggag agctgcacgc cattctgcgg cggcaggaag
     3001 atttttaccc attcctgaag gacaaccggg aaaagatcga gaagatcctg accttccgca
     3061 tcccctacta cgtgggccct ctggccaggg gaaacagcag attcgcctgg atgaccagaa
     3121 agagcgagga aaccatcacc ccctggaact tcgaggaagt ggtggacaag ggcgcttccg
     3181 cccagagctt catcgagcgg atgaccaact tcgataagaa cctgcccaac gagaaggtgc
     3241 tgcccaagca cagcctgctg tacgagtact tcaccgtgta taacgagctg accaaagtga
     3301 aatacgtgac cgagggaatg agaaagcccg ccttcctgag cggcgagcag aaaaaggcca
     3361 tcgtggacct gctgttcaag accaaccgga aagtgaccgt gaagcagctg aaagaggact
     3421 acttcaagaa aatcgagtgc ttcgactccg tggaaatctc cggcgtggaa gatcggttca
     3481 acgcctccct gggcacatac cacgatctgc tgaaaattat caaggacaag gacttcctgg
     3541 acaatgagga aaacgaggac attctggaag atatcgtgct gaccctgaca ctgtttgagg
     3601 acagagagat gatcgaggaa cggctgaaaa cctatgccca cctgttcgac gacaaagtga
     3661 tgaagcagct gaagcggcgg agatacaccg gctggggcag gctgagccgg aagctgatca
     3721 acggcatccg ggacaagcag tccggcaaga caatcctgga tttcctgaag tccgacggct
     3781 tcgccaacag aaacttcatg cagctgatcc acgacgacag cctgaccttt aaagaggaca
     3841 tccagaaagc ccaggtgtcc ggccagggcg atagcctgca cgagcacatt gccaatctgg
     3901 ccggcagccc cgccattaag aagggcatcc tgcagacagt gaaggtggtg gacgagctcg
     3961 tgaaagtgat gggccggcac aagcccgaga acatcgtgat cgaaatggcc agagagaacc
     4021 agaccaccca gaagggacag aagaacagcc gcgagagaat gaagcggatc gaagagggca
     4081 tcaaagagct gggcagccag atcctgaaag aacaccccgt ggaaaacacc cagctgcaga
     4141 acgagaagct gtacctgtac tacctgcaga atgggcggga tatgtacgtg gaccaggaac
     4201 tggacatcaa ccggctgtcc gactacgatg tggacgctat cgtgcctcag agctttctga
     4261 aggacgactc catcgacaac aaggtgctga ccagaagcga caagaaccgg ggcaagagcg
     4321 acaacgtgcc ctccgaagag gtcgtgaaga agatgaagaa ctactggcgg cagctgctga
     4381 acgccaagct gattacccag agaaagttcg acaatctgac caaggccgag agaggcggcc
     4441 tgagcgaact ggataaggcc ggcttcatca agagacagct ggtggaaacc cggcagatca
     4501 caaagcacgt ggcacagatc ctggactccc ggatgaacac taagtacgac gagaatgaca
     4561 agctgatccg ggaagtgaaa gtgatcaccc tgaagtccaa gctggtgtcc gatttccgga
     4621 aggatttcca gttttacaaa gtgcgcgaga tcaacaacta ccaccacgcc cacgacgcct
     4681 acctgaacgc cgtcgtggga accgccctga tcaaaaagta ccctaagctg gaaagcgagt
     4741 tcgtgtacgg cgactacaag gtgtacgacg tgcggaagat gatcgccaag agcgagcagg
     4801 aaatcggcaa ggctaccgcc aagtacttct tctacagcaa catcatgaac tttttcaaga
     4861 ccgagattac cctggccaac ggcgagatcc ggaagcggcc tctgatcgag acaaacggcg
     4921 aaaccgggga gatcgtgtgg gataagggcc gggattttgc caccgtgcgg aaagtgctga
     4981 gcatgcccca agtgaatatc gtgaaaaaga ccgaggtgca gacaggcggc ttcagcaaag
     5041 agtctatcct gcccaagagg aacagcgata agctgatcgc cagaaagaag gactgggacc
     5101 ctaagaagta cggcggcttc gacagcccca ccgtggccta ttctgtgctg gtggtggcca
     5161 aagtggaaaa gggcaagtcc aagaaactga agagtgtgaa agagctgctg gggatcacca
     5221 tcatggaaag aagcagcttc gagaagaatc ccatcgactt tctggaagcc aagggctaca
     5281 aagaagtgaa aaaggacctg atcatcaagc tgcctaagta ctccctgttc gagctggaaa
     5341 acggccggaa gagaatgctg gcctctgccg gcgaactgca gaagggaaac gaactggccc
     5401 tgccctccaa atatgtgaac ttcctgtacc tggccagcca ctatgagaag ctgaagggct
     5461 cccccgagga taatgagcag aaacagctgt ttgtggaaca gcacaagcac tacctggacg
     5521 agatcatcga gcagatcagc gagttctcca agagagtgat cctggccgac gctaatctgg
     5581 acaaagtgct gtccgcctac aacaagcacc gggataagcc catcagagag caggccgaga
     5641 atatcatcca cctgtttacc ctgaccaatc tgggagcccc tgccgccttc aagtactttg
     5701 acaccaccat cgaccggaag aggtacacca gcaccaaaga ggtgctggac gccaccctga
     5761 tccaccagag catcaccggc ctgtacgaga cacggatcga cctgtctcag ctgggaggtg
     5821 actctggagg atctagcgga ggatcctctg gcagcgagac accaggaaca agcgagtcag
     5881 caacaccaga gagcagtggc ggcagcagcg gcggcagcag caccctaaat atagaagatg
     5941 agtatcggct acatgagacc tcaaaagagc cagatgtttc tctagggtcc acatggctgt
     6001 ctgattttcc tcaggcctgg gcggaaaccg ggggcatggg actggcagtt cgccaagctc
     6061 ctctgatcat acctctgaaa gcaacctcta cccccgtgtc cataaaacaa taccccatgt
     6121 cacaagaagc cagactgggg atcaagcccc acatacagag actgttggac cagggaatac
     6181 tggtaccctg ccagtccccc tggaacacgc ccctgctacc cgttaagaaa ccagggacta
     6241 atgattatag gcctgtccag gatctgagag aagtcaacaa gcgggtggaa gacatccacc
     6301 ccaccgtgcc caacccttac aacctcttga gcgggctccc accgtcccac cagtggtaca
     6361 ctgtgcttga tttaaaggat gcctttttct gcctgagact ccaccccacc agtcagcctc
     6421 tcttcgcctt tgagtggaga gatccagaga tgggaatctc aggacaattg acctggacca
     6481 gactcccaca gggtttcaaa aacagtccca ccctgtttaa tgaggcactg cacagagacc
     6541 tagcagactt ccggatccag cacccagact tgatcctgct acagtacgtg gatgacttac
     6601 tgctggccgc cacttctgag ctagactgcc aacaaggtac tcgggccctg ttacaaaccc
     6661 tagggaacct cgggtatcgg gcctcggcca agaaagccca aatttgccag aaacaggtca
     6721 agtatctggg gtatcttcta aaagagggtc agagatggct gactgaggcc agaaaagaga
     6781 ctgtgatggg gcagcctact ccgaagaccc ctcgacaact aagggagttc ctagggaagg
     6841 caggcttctg tcgcctcttc atccctgggt ttgcagaaat ggcagccccc ctgtaccctc
     6901 tcaccaaacc ggggactctg tttaattggg gcccagacca acaaaaggcc tatcaagaaa
     6961 tcaagcaagc tcttctaact gccccagccc tggggttgcc agatttgact aagccctttg
     7021 aactctttgt cgacgagaag cagggctacg ccaaaggtgt cctaacgcaa aaactgggac
     7081 cttggcgtcg gccggtggcc tacctgtcca aaaagctaga cccagtagca gctgggtggc
     7141 ccccttgcct acggatggta gcagccattg ccgtactgac aaaggatgca ggcaagctaa
     7201 ccatgggaca gccactagtc attctggccc cccatgcagt agaggcacta gtcaaacaac
     7261 cccccgaccg ctggctttcc aacgcccgga tgactcacta tcaggccttg cttttggaca
     7321 cggaccgggt ccagttcgga ccggtggtag ccctgaaccc ggctacgctg ctcccactgc
     7381 ctgaggaagg gctgcaacac aactgccttg atatcctggc cgaagcccac ggaacccgac
     7441 ccgacctaac ggaccagccg ctcccagacg ccgaccacac ctggtacacg gatggaagca
     7501 gtctcttaca agagggacag cgtaaggcgg gagctgcggt gaccaccgag accgaggtaa
     7561 tctgggctaa agccctgcca gccgggacat ccgctcagcg ggctgaactg atagcactca
     7621 cccaggccct aaagatggca gaaggtaaga agctaaatgt ttatactgat agccgttatg
     7681 cttttgctac tgcccatatc catggagaaa tatacagaag gcgtgggtgg ctcacatcag
     7741 aaggcaaaga gatcaaaaat aaagacgaga tcttggccct actaaaagcc ctctttctgc
     7801 ccaaaagact tagcataatc cattgtccag gacatcaaaa gggacacagc gccgaggcta
     7861 gaggcaaccg gatggctgac caagcggccc gaaaggcagc catcacagag actccagaca
     7921 cctctaccct cctcatagaa aattcatcac cctctggcgg ctcaaaaaga accgccgacg
     7981 gcagcgaatt cgagcccaag aagaagagga aagtcggatc cggcgcaaca aacttctctc
     8041 tgctgaaaca agccggagat gtcgaagaga atcctggacc gaccgagtac aagcccacgg
     8101 tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg caccctcgcc gccgcgttcg
     8161 ccgactaccc cgccacgcgc cacaccgtcg atccggaccg ccacatcgag cgggtcaccg
     8221 agctgcaaga actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg tgggtcgcgg
     8281 acgacggcgc cgcggtggcg gtctggacca cgccggagag cgtcgaagcg ggggcggtgt
     8341 tcgccgagat cggcccgcgc atggccgagt tgagcggttc ccggctggcc gcgcagcaac
     8401 agatggaagg cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc ctggccaccg
     8461 tcggcgtgtc gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg ctccccggag
     8521 tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga gacctccgcg ccccgcaacc
     8581 tccccttcta cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg cccgaaggac
     8641 cgcgcacctg gtgcatgacc cgcaagcccg gtgccggatc catgcccacg ctactgcggg
     8701 tttatataga cggtcctcac gggatgggga aaaccaccac cacgcaactg ctggtggccc
     8761 tgggttcgcg cgacgatatc gtctacgtac ccgagccgat gacttactgg caggtgctgg
     8821 gggcttccga gacaatcgcg aacatctaca ccacacaaca ccgcctcgac cagggtgaga
     8881 tatcggccgg ggacgcggcg gtggtaatga caagcgccca gataacaatg ggcatgcctt
     8941 atgccgtgac cgacgccgtt ctggctcctc atatcggggg ggaggctggg agctcacatg
     9001 ccccgccccc ggccctcacc ctcatcttcg accgccatcc catcgccgcc ctcctgtgct
     9061 acccggccgc gcgatacctt atgggcagca tgacccccca ggccgtgctg gcgttcgtgg
     9121 ccctcatccc gccgaccttg cccggcacaa acatcgtgtt gggggccctt ccggaggaca
     9181 gacacatcga ccgcctggcc aaacgccagc gccccggcga gcggcttgac ctggctatgc
     9241 tggccgcgat tcgccgcgtt tacgggctgc ttgccaatac ggtgcggtat ctgcagggcg
     9301 gcgggtcgtg gcgggaggat tggggacagc tttcggggac ggccgtgccg ccccagggtg
     9361 ccgagcccca gagcaacgcg ggcccacgac cccatatcgg ggacacgtta tttaccctgt
     9421 ttcgggcccc cgagttgctg gcccccaacg gcgacctgta caacgtgttt gcctgggcct
     9481 tggacgtctt ggccaaacgc ctccgtccca tgcacgtctt tatcctggat tacgaccaat
     9541 cgcccgccgg ctgccgggac gccctgctgc aacttacctc cgggatggtc cagacccacg
     9601 tcaccacccc cggctccata ccgacgatct gcgacctggc gcgcacgttt gcccgggaga
     9661 tgggggaggc taactgacat atgtagatcc cccgggggat cagcctcgac tgtgccttct
     9721 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     9781 actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt
     9841 cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagacaat
     9901 agcaggcatg ctggggatgc ggtgggctct atgggcggcc gctctagaac tagtggatcc
     9961 cccgggtcac cattatcgtt tcagacccac ctcccaaccc cgaggggacc cagagagggc
    10021 ctatttccca tgattccttc atatttgcat atacgataca aggctgttag agagataatt
    10081 agaattaatt tgactgtaaa cacaaagata ttagtacaaa atacgtgacg tagaaagtaa
    10141 taatttcttg ggtagtttgc agttttaaaa ttatgtttta aaatggacta tcatatgctt
    10201 accgtaactt gaaagtattt cgatttcttg gctttatata tcttgtggaa aggacgaaac
    10261 acccgagacg gtagtcaaaa gcctccggtc ggaggctttt gacttggctg aggagtgcca
    10321 ttaagcaccg gtggagtgac gaccttcagc acgttcgtac tgttcaacga tggtgtagtc
    10381 ttcgttgtgg gaggtgatgt ccagtttgat gtcggttttg taagcacccg gcagctgaac
    10441 cggtttttta gccatgtagg tggttttaac ttcagcgtcg tagtgaccac cgtctttcag
    10501 tttcagacgc attttgattt cacctttcag agcaccgtct tccgggtaca tacgttcggt
    10561 ggaagcttcc caacccatgg tttttttctg cataaccgga ccgtcggacg ggaagttggt
    10621 accacgcagt ttaactttgt agatgaactc accgtcttgc agggaggagt cctgggtaac
    10681 ggtaacaaca ccaccgtctt cgaagttcat aacacgttcc catttgaaac cttccgggaa
    10741 ggacagtttc aggtagtccg ggatgtcagc cgggtgttta acgtaagctt tggaaccgta
    10801 ctggaactgc ggggacagga tgtcccaagc gaacggcagc ggaccacctt tggtaacttt
    10861 cagtttagcg gtctgggtac cttcgtacgg acgaccttca ccttcacctt cgatttcgaa
    10921 ctcgtgaccg ttaacggaac cttccatacg aactttgaaa cgcatgaact ctttgataac
    10981 gtcttcgcta ctcgccatag atcctttctc ctctttcaga tccgtgctca gtatctctat
    11041 cactgatagg gatgtcaatc tctatcactg atagggacgt ctcgtttttt tgggctgcag
    11101 gaattcgata aaagttttgt tactttatag aagaaatttt gagtttttgt tttttttaat
    11161 aaataaataa acataaataa attgtttgtt gaatttatta ttagtatgta agtgtaaata
    11221 taataaaact taatatctat tcaaattaat aaataaacct cgatatacag accgataaaa
    11281 cacatgcgtc aattttacac atgattatct ttaacgtacg tcacaatatg attatctttc
    11341 tagggttaat ctagctgcgt gttctgcagc gtgtcgagca tcttcatctg ctccatcacg
    11401 ctgtaaaaca catttgcacc gcgagtctgc ccgtcctcca cgggttcaaa aacgtgaatg
    11461 aacgaggcgc gctcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt
    11521 acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag
    11581 gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg ggacgcgccc
    11641 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
    11701 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
    11761 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
    11821 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc
    11881 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
    11941 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt
    12001 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
    12061 tttaacaaaa tattaacgct tacaatttag gtggcacttt tcggggaaat gtgcgcggaa
    12121 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
    12181 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
    12241 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
    12301 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
    12361 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
    12421 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
    12481 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
    12541 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
    12601 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
    12661 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
    12721 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
    12781 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
    12841 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
    12901 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
    12961 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
    13021 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
    13081 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
    13141 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
    13201 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
    13261 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
    13321 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
    13381 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
    13441 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
    13501 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
    13561 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
    13621 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
    13681 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
    13741 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
    13801 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
    13861 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
    13921 attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa
    13981 cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcccaata cgcaaacc
//
