LOCUS       sequence-441213-       13431 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Piggybac vector with doxycyclin-inducible prime editor for mammalian
            cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13431)
  AUTHORS   Koeppel J, Weller J, Peets EM, Pallaseni A, Kuzmin I, Raudvere U, 
            Peterson H, Liberante FG, Parts L
  TITLE     Prediction of prime editing insertion efficiencies using sequence 
            features and DNA repair determinants.
  JOURNAL   Nat Biotechnol. 2023 Feb 16. doi: 10.1038/s41587-023-01678-y.
  PUBMED    36797492
REFERENCE   2  (bases 1 to 13431)
  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. 2023 Feb 16. doi: 10.1038/s41587-023-01678-y."
COMMENT     Sequence Label: pPB-TREG3G-PE2-rtTA3G-P2A-eGFP
FEATURES             Location/Qualifiers
     source          1..13431
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             3888..3908
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    3941..4062
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3978..3997)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4032..4051
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4111..4127)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    4135..4151
                     /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(4159..4165,4166..4183,4184..4189))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4204..4225
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     4651..4670
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     polyA_signal    4669..4724
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     4770..4789
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     CDS             complement(4886..4906)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(join(4919..5629,5630..5632,5633..5635))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(4919..5629,5630..5632,5633..5635))
                     /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(4961..4982)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     5310..5329
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     CDS             complement(5636..5692)
                     /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             complement(5702..6445)
                     /codon_start=1
                     /product="modified rtTA protein that binds tightly to 
                     promoters containing the tet operator in the presence of 
                     doxycycline"
                     /label=Tet-On(R) 3G
                     /translation="MSRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHSCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLKQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     promoter        complement(6478..6978)
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     complement(6541..6559)
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     repeat_region   7449..7483
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(7522..7538)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    7546..7562
                     /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(7570..7576,7577..7594,7595..7600))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7615..7636
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(7924..8512)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8004..8023)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(8683..9474,9475..9543))
                     /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     9306..9325
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9544..9648)
                     /gene="bla"
                     /label=AmpR promoter
     repeat_region   9695..9757
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        10629..11006
                     /label=TRE3G promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G"
     protein_bind    10636..10654
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    10672..10690
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    10708..10726
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    10744..10762
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    10780..10798
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    10816..10834
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    10852..10870
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     10915..10939
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             11025..11045
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(11046..13431,1..128,129..131,132..1715)
                     /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             join(11046..13431,1..1715)
                     /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"
ORIGIN
        1 aagaacaccc cgtggaaaac acccagctgc agaacgagaa gctgtacctg tactacctgc
       61 agaatgggcg ggatatgtac gtggaccagg aactggacat caaccggctg tccgactacg
      121 atgtggacgc tatcgtgcct cagagctttc tgaaggacga ctccatcgac aacaaggtgc
      181 tgaccagaag cgacaagaac cggggcaaga gcgacaacgt gccctccgaa gaggtcgtga
      241 agaagatgaa gaactactgg cggcagctgc tgaacgccaa gctgattacc cagagaaagt
      301 tcgacaatct gaccaaggcc gagagaggcg gcctgagcga actggataag gccggcttca
      361 tcaagagaca gctggtggaa acccggcaga tcacaaagca cgtggcacag atcctggact
      421 cccggatgaa cactaagtac gacgagaatg acaagctgat ccgggaagtg aaagtgatca
      481 ccctgaagtc caagctggtg tccgatttcc ggaaggattt ccagttttac aaagtgcgcg
      541 agatcaacaa ctaccaccac gcccacgacg cctacctgaa cgccgtcgtg ggaaccgccc
      601 tgatcaaaaa gtaccctaag ctggaaagcg agttcgtgta cggcgactac aaggtgtacg
      661 acgtgcggaa gatgatcgcc aagagcgagc aggaaatcgg caaggctacc gccaagtact
      721 tcttctacag caacatcatg aactttttca agaccgagat taccctggcc aacggcgaga
      781 tccggaagcg gcctctgatc gagacaaacg gcgaaaccgg ggagatcgtg tgggataagg
      841 gccgggattt tgccaccgtg cggaaagtgc tgagcatgcc ccaagtgaat atcgtgaaaa
      901 agaccgaggt gcagacaggc ggcttcagca aagagtctat cctgcccaag aggaacagcg
      961 ataagctgat cgccagaaag aaggactggg accctaagaa gtacggcggc ttcgacagcc
     1021 ccaccgtggc ctattctgtg ctggtggtgg ccaaagtgga aaagggcaag tccaagaaac
     1081 tgaagagtgt gaaagagctg ctggggatca ccatcatgga aagaagcagc ttcgagaaga
     1141 atcccatcga ctttctggaa gccaagggct acaaagaagt gaaaaaggac ctgatcatca
     1201 agctgcctaa gtactccctg ttcgagctgg aaaacggccg gaagagaatg ctggcctctg
     1261 ccggcgaact gcagaaggga aacgaactgg ccctgccctc caaatatgtg aacttcctgt
     1321 acctggccag ccactatgag aagctgaagg gctcccccga ggataatgag cagaaacagc
     1381 tgtttgtgga acagcacaag cactacctgg acgagatcat cgagcagatc agcgagttct
     1441 ccaagagagt gatcctggcc gacgctaatc tggacaaagt gctgtccgcc tacaacaagc
     1501 accgggataa gcccatcaga gagcaggccg agaatatcat ccacctgttt accctgacca
     1561 atctgggagc ccctgccgcc ttcaagtact ttgacaccac catcgaccgg aagaggtaca
     1621 ccagcaccaa agaggtgctg gacgccaccc tgatccacca gagcatcacc ggcctgtacg
     1681 agacacggat cgacctgtct cagctgggag gtgactctgg aggatctagc ggaggatcct
     1741 ctggcagcga gacaccagga acaagcgagt cagcaacacc agagagcagt ggcggcagca
     1801 gcggcggcag cagcacccta aatatagaag atgagtatcg gctacatgag acctcaaaag
     1861 agccagatgt ttctctaggg tccacatggc tgtctgattt tcctcaggcc tgggcggaaa
     1921 ccgggggcat gggactggca gttcgccaag ctcctctgat catacctctg aaagcaacct
     1981 ctacccccgt gtccataaaa caatacccca tgtcacaaga agccagactg gggatcaagc
     2041 cccacataca gagactgttg gaccagggaa tactggtacc ctgccagtcc ccctggaaca
     2101 cgcccctgct acccgttaag aaaccaggga ctaatgatta taggcctgtc caggatctga
     2161 gagaagtcaa caagcgggtg gaagacatcc accccaccgt gcccaaccct tacaacctct
     2221 tgagcgggct cccaccgtcc caccagtggt acactgtgct tgatttaaag gatgcctttt
     2281 tctgcctgag actccacccc accagtcagc ctctcttcgc ctttgagtgg agagatccag
     2341 agatgggaat ctcaggacaa ttgacctgga ccagactccc acagggtttc aaaaacagtc
     2401 ccaccctgtt taatgaggca ctgcacagag acctagcaga cttccggatc cagcacccag
     2461 acttgatcct gctacagtac gtggatgact tactgctggc cgccacttct gagctagact
     2521 gccaacaagg tactcgggcc ctgttacaaa ccctagggaa cctcgggtat cgggcctcgg
     2581 ccaagaaagc ccaaatttgc cagaaacagg tcaagtatct ggggtatctt ctaaaagagg
     2641 gtcagagatg gctgactgag gccagaaaag agactgtgat ggggcagcct actccgaaga
     2701 cccctcgaca actaagggag ttcctaggga aggcaggctt ctgtcgcctc ttcatccctg
     2761 ggtttgcaga aatggcagcc cccctgtacc ctctcaccaa accggggact ctgtttaatt
     2821 ggggcccaga ccaacaaaag gcctatcaag aaatcaagca agctcttcta actgccccag
     2881 ccctggggtt gccagatttg actaagccct ttgaactctt tgtcgacgag aagcagggct
     2941 acgccaaagg tgtcctaacg caaaaactgg gaccttggcg tcggccggtg gcctacctgt
     3001 ccaaaaagct agacccagta gcagctgggt ggcccccttg cctacggatg gtagcagcca
     3061 ttgccgtact gacaaaggat gcaggcaagc taaccatggg acagccacta gtcattctgg
     3121 ccccccatgc agtagaggca ctagtcaaac aaccccccga ccgctggctt tccaacgccc
     3181 ggatgactca ctatcaggcc ttgcttttgg acacggaccg ggtccagttc ggaccggtgg
     3241 tagccctgaa cccggctacg ctgctcccac tgcctgagga agggctgcaa cacaactgcc
     3301 ttgatatcct ggccgaagcc cacggaaccc gacccgacct aacggaccag ccgctcccag
     3361 acgccgacca cacctggtac acggatggaa gcagtctctt acaagaggga cagcgtaagg
     3421 cgggagctgc ggtgaccacc gagaccgagg taatctgggc taaagccctg ccagccggga
     3481 catccgctca gcgggctgaa ctgatagcac tcacccaggc cctaaagatg gcagaaggta
     3541 agaagctaaa tgtttatact gatagccgtt atgcttttgc tactgcccat atccatggag
     3601 aaatatacag aaggcgtggg tggctcacat cagaaggcaa agagatcaaa aataaagacg
     3661 agatcttggc cctactaaaa gccctctttc tgcccaaaag acttagcata atccattgtc
     3721 caggacatca aaagggacac agcgccgagg ctagaggcaa ccggatggct gaccaagcgg
     3781 cccgaaaggc agccatcaca gagactccag acacctctac cctcctcata gaaaattcat
     3841 caccctctgg cggctcaaaa agaaccgccg acggcagcga attcgagccc aagaagaaga
     3901 ggaaagtctg ataaaaactc gcgaatcgat cgcccacccc aacttgttta ttgcagctta
     3961 taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat ttttttcact
     4021 gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct gtataccgtc
     4081 gacctctagc tagagcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta
     4141 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     4201 ctaatgagtg agctaactca cattaattgc gttgcgctca ctgcccgctt tccagtcggg
     4261 aaacctgtcg tgccagctgc attaatgaat cggccaacgc gcggggagag gcggtttgcg
     4321 tattgggcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg
     4381 gcgagcggta tcagctcact caaaggacgg ccgcacatgt aaggaaaatt ttagggatgt
     4441 taaagaaaaa aataacacaa aacaaaatat aaaaaaaatc taacctcaag tcaaggcttt
     4501 tctatggaat aaggaatgga cagcaggggg ctgtttcata tactgatgac ctctttatag
     4561 ccaacctttg ttcatggcag ccagcatatg ggcatatgtt gccaaactct aaaccaaata
     4621 ctcattctga tgttttaaat gatttgccct cccatatgtc cttccgagtg agagacacaa
     4681 aaaattccaa cacactattg caatgaaaat aaatttcctt tattagccag aagtcagatg
     4741 ctcaaggggc ttcatgatgt ccccataatt tttggcagag ggaaaaagat ctcagtggta
     4801 tttgtgagcc agggcattgg ccacaccagt caccaccttc tgataggcag cctgcacctg
     4861 aggagtgaat tcacgcgttt tattagactt tcctcttctt cttgggagaa ccaccagact
     4921 tgtacagctc gtccatgccg agagtgatcc cggcggcggt cacgaactcc agcaggacca
     4981 tgtgatcgcg cttctcgttg gggtctttgc tcagggcgga ctgggtgctc aggtagtggt
     5041 tgtcgggcag cagcacgggg ccgtcgccga tgggggtgtt ctgctggtag tggtcggcga
     5101 gctgcacgct gccgtcctcg atgttgtggc ggatcttgaa gttcaccttg atgccgttct
     5161 tctgcttgtc ggccatgata tagacgttgt ggctgttgta gttgtactcc agcttgtgcc
     5221 ccaggatgtt gccgtcctcc ttgaagtcga tgcccttcag ctcgatgcgg ttcaccaggg
     5281 tgtcgccctc gaacttcacc tcggcgcggg tcttgtagtt gccgtcgtcc ttgaagaaga
     5341 tggtgcgctc ctggacgtag ccttcgggca tggcggactt gaagaagtcg tgctgcttca
     5401 tgtggtcggg gtagcggctg aagcactgca ctccataggt cagggtggtc accagcgtgg
     5461 gccagggcac gggcagcttg ccggtggtgc agatgaactt cagggtcagc ttgccgtagg
     5521 tggcatcgcc ctcgccctcg ccggacacgc tgaacttgtg gccgtttacg tcgccgtcca
     5581 gctcgaccag gatgggcacc accccggtga acagctcctc gcccttgctc accataggtc
     5641 cagggttctc ctccacgtct ccagcctgct tcagcaggct gaagttagta gctccgcttc
     5701 cgcctggcag catatcgaga tcgaagtcat cgagagcgtc agcagggagc atatccagat
     5761 caaaatcatc cagggcatcg gcgggcagca tgtccaggtc gaagtcgtcc agggcgtcgg
     5821 tgggtccgcc gctctcgcac ttcagctgct tctcgaggcc gcagatgatc agttccaggc
     5881 cgaacaggaa ggcgggctcg gcgccctgtc tgtcgaacag ctcgatggcc tgcttcagca
     5941 gggggggcat gctgtcggtg gtgggtgtct ctctttcctc tttggcgacc tggtgttcct
     6001 gttcttccag cacgcagccc agggtgaagt ggcccacggc gctcagggcg tacagggcgt
     6061 tttccaggct gaagccctgc tggcacagga aggccagctg gttttccagt gtctcgtact
     6121 gcttctcggt gggtctggtg cccaggtgca ctttggcgcc gtcccggtgg ctcagcaggg
     6181 cgcatctgta gctcttggcg ttgttccgca ggaagtcctg ccagctctcg ccttccagag
     6241 ggcagctgtg ggtgtggtgc cggtccagca tctcgatggg cagggcgtcc agcagggccc
     6301 gcttgttctt cacgtgccag tacagggtgg gctgttccac gcccagtttc tgggccagct
     6361 tccgggtggt caggccctcg atgcccacgc cgttcagcag ttccagggcg ctgttgatca
     6421 ctttgctctt gtccagccgg ctcatggtgg ctattccgga gctagcggta tcgatccctg
     6481 gggagagagg tcggtgattc ggtcaacgag ggagccgact gccgacgtgc gctccggagg
     6541 cttgcagaat gcggaacacc gcgcgggcag gaacagggcc cacactaccg ccccacaccc
     6601 cgcctcccgc accgcccctt cccggccgct gctctcggcg cgccctgctg agcagccgct
     6661 attggccaca gcccatcgcg gtcggcgcgc tgccattgct ccctggcgct gtccgtctgc
     6721 gagggtacta gtgagtcgtg cggcttccgt ttgtcacgtc cggcacgccg cgaaccgcaa
     6781 ggaaccttcc cgacttaggg gcggagcagg aagcgtcgcc ggggggccca caagggtagc
     6841 ggcgaagatc cgggtgacgc tgcgaacgga cgtgaagaat gtgcgagacc cagggtcggc
     6901 gccgctgcgt ttcccggaac cacgcccaga gcagccgcgt ccctgcgcaa acccagggct
     6961 gccttggaaa aggcgcaagg gtcccctcgg ggttgggagg tgggtctgaa acgataatgg
     7021 tgaatatccc tgcctaactc tattcactat agaaagtaca gcaaaaacta ttccaattgc
     7081 attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttgttt aaactggtaa
     7141 gctcgtaccg tgatacatgc ggcaggccgt taatgatatc tataacaaga aaatatatat
     7201 ataataagtt atcacgtaag tagaacatga aataacaata taattatcgt atgagttaaa
     7261 tcttaaaagt cacgtaaaag ataatcatgc gtcattttga ctcacgcggt cgttatagtt
     7321 caaaatcagt gacacttacc gcattgacaa gcacgcctca cgggagctcc aagcggcgac
     7381 tgagatgtcc taaatgcaca gcgacggatt cgcgctattt agaaagagag agcaatattt
     7441 caagaatgca tgcgtcaatt ttacgcagac tatctttcta gggttaaaca cgacggctca
     7501 cagccggtaa gagtaacccg ggtcatagct gtttcctgtg tgaaattgtt atccgctcac
     7561 aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt
     7621 gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc
     7681 gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg
     7741 ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt
     7801 atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa
     7861 gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
     7921 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     7981 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     8041 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     8101 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     8161 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     8221 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     8281 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     8341 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     8401 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     8461 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     8521 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     8581 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     8641 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     8701 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     8761 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     8821 gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     8881 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     8941 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     9001 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     9061 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     9121 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     9181 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     9241 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     9301 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     9361 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     9421 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     9481 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     9541 catactcttc ctttttcaat attattgaag catttatcag ggttattgta tcatgagcgg
     9601 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     9661 aaaagtgcca cctgacgtct aagaaaccgg ttaaccctag aaagataatc atattgtgac
     9721 gtacgttaaa gataatcatg cgtaaaattg acgcatgtgt tttatcggtc tgtatatcga
     9781 ggtttattta ttaatttgaa tagatattaa gttttattat atttacactt acatactaat
     9841 aataaattca acaaacaatt tatttatgtt tatttattta ttaaaaaaaa acaaaaactc
     9901 aaaatttctt ctataaagta acaaagcaaa aacacgttag ggccttaatt aagggaggtg
     9961 gtccctgcag ttacgccaat gataaccccc gccagaaaaa tcttagtagc cttccctttt
    10021 tgttttccgt gccccaactc ggcggattga ctcggcccct tccggaaaca cccgaatcaa
    10081 cttctagtca aattattgtt cacgccgcaa tgacccaccc ctggcccgcg tctgtggaac
    10141 tgacccctgg tgtacaggag agttcgctgc tgaaagtggt cccaaagggg tactagtttt
    10201 taagctccca actccccctc ccccagcgtc tggaggattc cacaccctcg caccgcaggg
    10261 gcgaggaagt gggcggagtc cggttttggc gccagccgct gaggctgcca agcagaaaag
    10321 ccaccgctga ggagactccg gtcactgtcc tcgccccgcc tcccccttcc ctccccttgg
    10381 ggaccaccgg gcgccacgcc gcgaacggta agtgccgcgg tcgtcggcgc ctccgccctc
    10441 cccctagggc cccaattccc agcgggcgcg gcgcgcggcc cctccccccg ccgggcgcgc
    10501 gcccgctgcc ccgcccttcg tggccgcccg gcgtgggcgg tgccacccct ccccccggcg
    10561 gccccgcgcg cagctcccgg ctccctcccc cttcggatgt ggcttgagct gtaggcgcgg
    10621 agggtaccag tttactccct atcagtgata gagaacgtat gaagagttta ctccctatca
    10681 gtgatagaga acgtatgcag actttactcc ctatcagtga tagagaacgt ataaggagtt
    10741 tactccctat cagtgataga gaacgtatga ccagtttact ccctatcagt gatagagaac
    10801 gtatctacag tttactccct atcagtgata gagaacgtat atccagttta ctccctatca
    10861 gtgatagaga acgtataagc tttaggcgtg tacggtgggc gcctataaaa gcagagctcg
    10921 tttagtgaac cgtcagatcg cctggagcaa ttccacaaca cttttgtctt ataccaactt
    10981 tccgtaccac ttcctaccct cgtaaaggat ccgccgccac catgccaaag aagaagcgga
    11041 aagtcgacaa gaagtacagc atcggcctgg acatcggcac caactctgtg ggctgggccg
    11101 tgatcaccga cgagtacaag gtgcccagca agaaattcaa ggtgctgggc aacaccgacc
    11161 ggcacagcat caagaagaac ctgatcggag ccctgctgtt cgacagcggc gaaacagccg
    11221 aggccacccg gctgaagaga accgccagaa gaagatacac cagacggaag aaccggatct
    11281 gctatctgca agagatcttc agcaacgaga tggccaaggt ggacgacagc ttcttccaca
    11341 gactggaaga gtccttcctg gtggaagagg ataagaagca cgagcggcac cccatcttcg
    11401 gcaacatcgt ggacgaggtg gcctaccacg agaagtaccc caccatctac cacctgagaa
    11461 agaaactggt ggacagcacc gacaaggccg acctgcggct gatctatctg gccctggccc
    11521 acatgatcaa gttccggggc cacttcctga tcgagggcga cctgaacccc gacaacagcg
    11581 acgtggacaa gctgttcatc cagctggtgc agacctacaa ccagctgttc gaggaaaacc
    11641 ccatcaacgc cagcggcgtg gacgccaagg ccatcctgtc tgccagactg agcaagagca
    11701 gacggctgga aaatctgatc gcccagctgc ccggcgagaa gaagaatggc ctgttcggaa
    11761 acctgattgc cctgagcctg ggcctgaccc ccaacttcaa gagcaacttc gacctggccg
    11821 aggatgccaa actgcagctg agcaaggaca cctacgacga cgacctggac aacctgctgg
    11881 cccagatcgg cgaccagtac gccgacctgt ttctggccgc caagaacctg tccgacgcca
    11941 tcctgctgag cgacatcctg agagtgaaca ccgagatcac caaggccccc ctgagcgcct
    12001 ctatgatcaa gagatacgac gagcaccacc aggacctgac cctgctgaaa gctctcgtgc
    12061 ggcagcagct gcctgagaag tacaaagaga ttttcttcga ccagagcaag aacggctacg
    12121 ccggctacat tgacggcgga gccagccagg aagagttcta caagttcatc aagcccatcc
    12181 tggaaaagat ggacggcacc gaggaactgc tcgtgaagct gaacagagag gacctgctgc
    12241 ggaagcagcg gaccttcgac aacggcagca tcccccacca gatccacctg ggagagctgc
    12301 acgccattct gcggcggcag gaagattttt acccattcct gaaggacaac cgggaaaaga
    12361 tcgagaagat cctgaccttc cgcatcccct actacgtggg ccctctggcc aggggaaaca
    12421 gcagattcgc ctggatgacc agaaagagcg aggaaaccat caccccctgg aacttcgagg
    12481 aagtggtgga caagggcgct tccgcccaga gcttcatcga gcggatgacc aacttcgata
    12541 agaacctgcc caacgagaag gtgctgccca agcacagcct gctgtacgag tacttcaccg
    12601 tgtataacga gctgaccaaa gtgaaatacg tgaccgaggg aatgagaaag cccgccttcc
    12661 tgagcggcga gcagaaaaag gccatcgtgg acctgctgtt caagaccaac cggaaagtga
    12721 ccgtgaagca gctgaaagag gactacttca agaaaatcga gtgcttcgac tccgtggaaa
    12781 tctccggcgt ggaagatcgg ttcaacgcct ccctgggcac ataccacgat ctgctgaaaa
    12841 ttatcaagga caaggacttc ctggacaatg aggaaaacga ggacattctg gaagatatcg
    12901 tgctgaccct gacactgttt gaggacagag agatgatcga ggaacggctg aaaacctatg
    12961 cccacctgtt cgacgacaaa gtgatgaagc agctgaagcg gcggagatac accggctggg
    13021 gcaggctgag ccggaagctg atcaacggca tccgggacaa gcagtccggc aagacaatcc
    13081 tggatttcct gaagtccgac ggcttcgcca acagaaactt catgcagctg atccacgacg
    13141 acagcctgac ctttaaagag gacatccaga aagcccaggt gtccggccag ggcgatagcc
    13201 tgcacgagca cattgccaat ctggccggca gccccgccat taagaagggc atcctgcaga
    13261 cagtgaaggt ggtggacgag ctcgtgaaag tgatgggccg gcacaagccc gagaacatcg
    13321 tgatcgaaat ggccagagag aaccagacca cccagaaggg acagaagaac agccgcgaga
    13381 gaatgaagcg gatcgaagag ggcatcaaag agctgggcag ccagatcctg a
//
