LOCUS       sequence-360164-       10827 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  GFP knock-in for beta3-tubulin-GFP (amino acid position: stop 
            codon).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10827)
  AUTHORS   Droogers WJ, Willems J, MacGillavry HD, de Jong APH
  TITLE     Duplex Labeling and Manipulation of Neuronal Proteins Using 
            Sequential CRISPR/Cas9 Gene Editing.
  JOURNAL   eNeuro. 2022 Jul 18;9(4):ENEURO.0056-22.2022. doi: 
            10.1523/ENEURO.0056-22.2022.
  PUBMED    35851300
REFERENCE   2  (bases 1 to 10827)
  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: 
            "10.1523/ENEURO.0056-22"; journalName: "eNeuro"; date: "2022-07-18- 
            18"; volume: "9"; issue: "4"; pages: "ENEURO.0056-22.2022"
COMMENT     Sequence Label: pORANGE Tubb3-3xGFP KI
FEATURES             Location/Qualifiers
     source          1..10827
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        39..287
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     39..59
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     210..229
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        307..382
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     CDS             join(509..511,512..1225)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(509..511,512..1222)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(1302..1304,1305..2018)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(1302..1304,1305..2015)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(2095..2097,2098..2811)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(2095..2097,2098..2808)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     enhancer        2869..3154
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        3156..3433
                     /label=chicken beta-actin promoter
     intron          3434..3661
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     CDS             3682..3708
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             3712..3732
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3742..7842
                     /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
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             3742..7842
                     /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
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             7846..7866
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(7906..7923)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    7912..8119
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   8128..8268
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   8128..8257
                     /label=AAV2 ITR
     rep_origin      8343..8798
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8430..8449)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     8640..8661
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(8815..8834)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8934..8956
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(8994..9012)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        9080..9184
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(9185..9253,9254..10045)
                     /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(9403..9422)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      10216..10804
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     10705..10724
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 acggttcctg gccttttgct ggccttttgc tcacatgtga gggcctattt cccatgattc
       61 cttcatattt gcatatacga tacaaggctg ttagagagat aattggaatt aatttgactg
      121 taaacacaaa gatattagta caaaatacgt gacgtagaaa gtaataattt cttgggtagt
      181 ttgcagtttt aaaattatgt tttaaaatgg actatcatat gcttaccgta acttgaaagt
      241 atttcgattt cttggcttta tatatcttgt ggaaaggacg aaacaccgct gcgagcaact
      301 tcacttgttt tagagctaga aatagcaagt taaaataagg ctagtccgtt atcaacttga
      361 aaaagtggca ccgagtcggt gcttttttgt tttagagcta gaaatagcaa gttaaaataa
      421 ggctagtccg tttttagcgc gtgcgccaat tctgcagaca aatggctcta gaagcttgct
      481 gcgagcaact tcacttgggg gatcaggcgt gagcaagggc gaggagctgt tcaccggggt
      541 ggtgcccatc ctggtcgagc tggacggcga cgtaaacggc cacaagttca gcgtgtccgg
      601 cgagggcgag ggcgatgcca cctacggcaa gctgaccctg aagttcatct gcaccaccgg
      661 caagctgccc gtgccctggc ccaccctcgt gaccaccctg acctacggcg tgcagtgctt
      721 cagccgctac cccgaccaca tgaagcagca cgacttcttc aagtccgcca tgcccgaagg
      781 ctacgtccag gagcgcacca tcttcttcaa ggacgacggc aactacaaga cccgcgccga
      841 ggtgaagttc gagggcgaca ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa
      901 ggaggacggc aacatcctgg ggcacaagct ggagtacaac tacaacagcc acaacgtcta
      961 tatcatggcc gacaagcaga agaacggcat caaggtgaac ttcaagatcc gccacaacat
     1021 cgaggacggc agcgtgcagc tcgccgacca ctaccagcag aacaccccca tcggcgacgg
     1081 ccccgtgctg ctgcccgaca accactacct gagcacccag tccgccctga gcaaagaccc
     1141 caacgagaag cgcgatcaca tggtcctgct ggagttcgtg accgccgccg ggatcactct
     1201 cggcatggac gagctgtaca agtaatgtgc tgcgagcaac ttcacttggg ctcgaggatc
     1261 ctagaagctt gctgcgagca acttcacttg ggggatcagg cgtgagcaag ggcgaggagc
     1321 tgttcaccgg ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt
     1381 tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca
     1441 tctgcaccac cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg
     1501 gcgtgcagtg cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg
     1561 ccatgcccga aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca
     1621 agacccgcgc cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg
     1681 gcatcgactt caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca
     1741 gccacaacgt ctatatcatg gccgacaagc agaagaacgg catcaaggtg aacttcaaga
     1801 tccgccacaa catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc
     1861 ccatcggcga cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccgccc
     1921 tgagcaaaga ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg
     1981 ccgggatcac tctcggcatg gacgagctgt acaagtaatg tgctgcgagc aacttcactt
     2041 gggctcgagg atcctagaag cttgctgcga gcaacttcac ttgggggatc aggcgtgagc
     2101 aagggcgagg agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta
     2161 aacggccaca agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg
     2221 accctgaagt tcatctgcac caccggcaag ctgcccgtgc cctggcccac cctcgtgacc
     2281 accctgacct acggcgtgca gtgcttcagc cgctaccccg accacatgaa gcagcacgac
     2341 ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc gcaccatctt cttcaaggac
     2401 gacggcaact acaagacccg cgccgaggtg aagttcgagg gcgacaccct ggtgaaccgc
     2461 atcgagctga agggcatcga cttcaaggag gacggcaaca tcctggggca caagctggag
     2521 tacaactaca acagccacaa cgtctatatc atggccgaca agcagaagaa cggcatcaag
     2581 gtgaacttca agatccgcca caacatcgag gacggcagcg tgcagctcgc cgaccactac
     2641 cagcagaaca cccccatcgg cgacggcccc gtgctgctgc ccgacaacca ctacctgagc
     2701 acccagtccg ccctgagcaa agaccccaac gagaagcgcg atcacatggt cctgctggag
     2761 ttcgtgaccg ccgccgggat cactctcggc atggacgagc tgtacaagta atgtgctgcg
     2821 agcaacttca cttgggctcg aggatcctag gacgcgtcca gaggtacccg ttacataact
     2881 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaatagt
     2941 aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt aaactgccca
     3001 cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg tcaatgacgg
     3061 taaatggccc gcctggcatt gtgcccagta catgacctta tgggactttc ctacttggca
     3121 gtacatctac gtattagtca tcgctattac catggtcgag gtgagcccca cgttctgctt
     3181 cactctcccc atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt
     3241 attttgtgca gcgatggggg cggggggggg gggggggcgc gcgccrggsg gggsggggsg
     3301 gggsgrgggg sggggsgggg cgrggcggrg aggkgcggcg gcagccaatc agagcggcgc
     3361 gctccgaaag tttcctttta tggcgaggcg gcggcggcgg cggccctata aaaagcgaag
     3421 cgcgcggcgg gcgggagtcg ctgcgcgctg ccttcgcccc gtgccccgct ccgccgccgc
     3481 ctcgcgccgc ccgccccggc tctgactgac cgcgttactc ccacaggtga gcgggcggga
     3541 cggcccttct cctccgggct gtaattagct gagcaagagg taagggttta agggatggtt
     3601 ggttggtggg gtattaatgt ttaattacct ggagcacctg cctgaaatca ctttttttca
     3661 ggttggaccg gtgccaccat gtacccatac gatgttccag attacgcttc gccgaagaaa
     3721 aagcgcaagg tcgaagcgtc cgacaagaag tacagcatcg gcctggacat cggcaccaac
     3781 tctgtgggct gggccgtgat caccgacgag tacaaggtgc ccagcaagaa attcaaggtg
     3841 ctgggcaaca ccgaccggca cagcatcaag aagaacctga tcggagccct gctgttcgac
     3901 agcggcgaaa cagccgaggc cacccggctg aagagaaccg ccagaagaag atacaccaga
     3961 cggaagaacc ggatctgcta tctgcaagag atcttcagca acgagatggc caaggtggac
     4021 gacagcttct tccacagact ggaagagtcc ttcctggtgg aagaggataa gaagcacgag
     4081 cggcacccca tcttcggcaa catcgtggac gaggtggcct accacgagaa gtaccccacc
     4141 atctaccacc tgagaaagaa actggtggac agcaccgaca aggccgacct gcggctgatc
     4201 tatctggccc tggcccacat gatcaagttc cggggccact tcctgatcga gggcgacctg
     4261 aaccccgaca acagcgacgt ggacaagctg ttcatccagc tggtgcagac ctacaaccag
     4321 ctgttcgagg aaaaccccat caacgccagc ggcgtggacg ccaaggccat cctgtctgcc
     4381 agactgagca agagcagacg gctggaaaat ctgatcgccc agctgcccgg cgagaagaag
     4441 aatggcctgt tcggaaacct gattgccctg agcctgggcc tgacccccaa cttcaagagc
     4501 aacttcgacc tggccgagga tgccaaactg cagctgagca aggacaccta cgacgacgac
     4561 ctggacaacc tgctggccca gatcggcgac cagtacgccg acctgtttct ggccgccaag
     4621 aacctgtccg acgccatcct gctgagcgac atcctgagag tgaacaccga gatcaccaag
     4681 gcccccctga gcgcctctat gatcaagaga tacgacgagc accaccagga cctgaccctg
     4741 ctgaaagctc tcgtgcggca gcagctgcct gagaagtaca aagagatttt cttcgaccag
     4801 agcaagaacg gctacgccgg ctacattgac ggcggagcca gccaggaaga gttctacaag
     4861 ttcatcaagc ccatcctgga aaagatggac ggcaccgagg aactgctcgt gaagctgaac
     4921 agagaggacc tgctgcggaa gcagcggacc ttcgacaacg gcagcatccc ccaccagatc
     4981 cacctgggag agctgcacgc cattctgcgg cggcaggaag atttttaccc attcctgaag
     5041 gacaaccggg aaaagatcga gaagatcctg accttccgca tcccctacta cgtgggccct
     5101 ctggccaggg gaaacagcag attcgcctgg atgaccagaa agagcgagga aaccatcacc
     5161 ccctggaact tcgaggaagt ggtggacaag ggcgcttccg cccagagctt catcgagcgg
     5221 atgaccaact tcgataagaa cctgcccaac gagaaggtgc tgcccaagca cagcctgctg
     5281 tacgagtact tcaccgtgta taacgagctg accaaagtga aatacgtgac cgagggaatg
     5341 agaaagcccg ccttcctgag cggcgagcag aaaaaggcca tcgtggacct gctgttcaag
     5401 accaaccgga aagtgaccgt gaagcagctg aaagaggact acttcaagaa aatcgagtgc
     5461 ttcgactccg tggaaatctc cggcgtggaa gatcggttca acgcctccct gggcacatac
     5521 cacgatctgc tgaaaattat caaggacaag gacttcctgg acaatgagga aaacgaggac
     5581 attctggaag atatcgtgct gaccctgaca ctgtttgagg acagagagat gatcgaggaa
     5641 cggctgaaaa cctatgccca cctgttcgac gacaaagtga tgaagcagct gaagcggcgg
     5701 agatacaccg gctggggcag gctgagccgg aagctgatca acggcatccg ggacaagcag
     5761 tccggcaaga caatcctgga tttcctgaag tccgacggct tcgccaacag aaacttcatg
     5821 cagctgatcc acgacgacag cctgaccttt aaagaggaca tccagaaagc ccaggtgtcc
     5881 ggccagggcg atagcctgca cgagcacatt gccaatctgg ccggcagccc cgccattaag
     5941 aagggcatcc tgcagacagt gaaggtggtg gacgagctcg tgaaagtgat gggccggcac
     6001 aagcccgaga acatcgtgat cgaaatggcc agagagaacc agaccaccca gaagggacag
     6061 aagaacagcc gcgagagaat gaagcggatc gaagagggca tcaaagagct gggcagccag
     6121 atcctgaaag aacaccccgt ggaaaacacc cagctgcaga acgagaagct gtacctgtac
     6181 tacctgcaga atgggcggga tatgtacgtg gaccaggaac tggacatcaa ccggctgtcc
     6241 gactacgatg tggaccatat cgtgcctcag agctttctga aggacgactc catcgacaac
     6301 aaggtgctga ccagaagcga caagaaccgg ggcaagagcg acaacgtgcc ctccgaagag
     6361 gtcgtgaaga agatgaagaa ctactggcgg cagctgctga acgccaagct gattacccag
     6421 agaaagttcg acaatctgac caaggccgag agaggcggcc tgagcgaact ggataaggcc
     6481 ggcttcatca agagacagct ggtggaaacc cggcagatca caaagcacgt ggcacagatc
     6541 ctggactccc ggatgaacac taagtacgac gagaatgaca agctgatccg ggaagtgaaa
     6601 gtgatcaccc tgaagtccaa gctggtgtcc gatttccgga aggatttcca gttttacaaa
     6661 gtgcgcgaga tcaacaacta ccaccacgcc cacgacgcct acctgaacgc cgtcgtggga
     6721 accgccctga tcaaaaagta ccctaagctg gaaagcgagt tcgtgtacgg cgactacaag
     6781 gtgtacgacg tgcggaagat gatcgccaag agcgagcagg aaatcggcaa ggctaccgcc
     6841 aagtacttct tctacagcaa catcatgaac tttttcaaga ccgagattac cctggccaac
     6901 ggcgagatcc ggaagcggcc tctgatcgag acaaacggcg aaaccgggga gatcgtgtgg
     6961 gataagggcc gggattttgc caccgtgcgg aaagtgctga gcatgcccca agtgaatatc
     7021 gtgaaaaaga ccgaggtgca gacaggcggc ttcagcaaag agtctatcct gcccaagagg
     7081 aacagcgata agctgatcgc cagaaagaag gactgggacc ctaagaagta cggcggcttc
     7141 gacagcccca ccgtggccta ttctgtgctg gtggtggcca aagtggaaaa gggcaagtcc
     7201 aagaaactga agagtgtgaa agagctgctg gggatcacca tcatggaaag aagcagcttc
     7261 gagaagaatc ccatcgactt tctggaagcc aagggctaca aagaagtgaa aaaggacctg
     7321 atcatcaagc tgcctaagta ctccctgttc gagctggaaa acggccggaa gagaatgctg
     7381 gcctctgccg gcgaactgca gaagggaaac gaactggccc tgccctccaa atatgtgaac
     7441 ttcctgtacc tggccagcca ctatgagaag ctgaagggct cccccgagga taatgagcag
     7501 aaacagctgt ttgtggaaca gcacaagcac tacctggacg agatcatcga gcagatcagc
     7561 gagttctcca agagagtgat cctggccgac gctaatctgg acaaagtgct gtccgcctac
     7621 aacaagcacc gggataagcc catcagagag caggccgaga atatcatcca cctgtttacc
     7681 ctgaccaatc tgggagcccc tgccgccttc aagtactttg acaccaccat cgaccggaag
     7741 aggtacacca gcaccaaaga ggtgctggac gccaccctga tccaccagag catcaccggc
     7801 ctgtacgaga cacggatcga cctgtctcag ctgggaggcg acagccccaa gaagaagaga
     7861 aaggtggagg ccagctaaga attctaacta gagctcgctg atcagcctcg actgtgcctt
     7921 ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg
     7981 ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt
     8041 gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaga
     8101 atagcaggca tgctggggag cggccgcagg aacccctagt gatggagttg gccactccct
     8161 ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct
     8221 ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg cctgcagggg cgcctgatgc
     8281 ggtattttct ccttacgcat ctgtgcggta tttcacaccg catacgtcaa agcaaccata
     8341 gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     8401 cgctacactt gccagcgcct tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     8461 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     8521 tagtgcttta cggcacctcg accccaaaaa acttgatttg ggtgatggtt cacgtagtgg
     8581 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     8641 tggactcttg ttccaaactg gaacaacact caactctatc tcgggctatt cttttgattt
     8701 ataagggatt ttgccgattt cggtctattg gttaaaaaat gagctgattt aacaaaaatt
     8761 taacgcgaat tttaacaaaa tattaacgtt tacaatttta tggtgcactc tcagtacaat
     8821 ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc
     8881 ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag
     8941 ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa agggcctcgt
     9001 gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg
     9061 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     9121 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     9181 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     9241 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     9301 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     9361 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     9421 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     9481 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     9541 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     9601 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     9661 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     9721 gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     9781 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     9841 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     9901 aagccgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     9961 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
    10021 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
    10081 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
    10141 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
    10201 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
    10261 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
    10321 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
    10381 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
    10441 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
    10501 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
    10561 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
    10621 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
    10681 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
    10741 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
    10801 gaaaaacgcc agcaacgcgg ccttttt
//
