LOCUS       sequence-382130-       13021 bp    DNA     circular SYN 14-APR-2025
DEFINITION  CRISPR vector for cloning sgDNAs to target genes of several 
            Aspergillus species (especially section Flavi).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13021)
  AUTHORS   Chang PK
  TITLE     A Simple CRISPR/Cas9 System for Efficiently Targeting Genes of 
            Aspergillus Section Flavi Species, Aspergillus nidulans, Aspergillus
            fumigatus, Aspergillus terreus, and Aspergillus niger.
  JOURNAL   Microbiol Spectr. 2023 Feb 14;11(1):e0464822. doi: 
            10.1128/spectrum.04648-22. Epub 2023 Jan 18.
  PUBMED    36651760
REFERENCE   2  (bases 1 to 13021)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Microbiol 
            Spectr."; date: "2023-02-14"; volume: "11(1):e0464822. doi"; pages: 
            " 10.1128/spectrum.04648-22. Epub 2023 Jan 18"
COMMENT     Sequence Label: pAsp-AMA-gpdA-ptr
FEATURES             Location/Qualifiers
     source          1..13021
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     3102..3123
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     gene            complement(3242..5249)
                     /product="pyrithiamine resistance gene of Aspergillus 
                     oryzae (Kubodera et al., 2000)"
                     /label=ptrA
                     /note="useful as a dominant selectable marker in 
                     Aspergillus"
     CDS             complement(join(3486..4132,4191..4527))
                     /codon_start=1
                     /gene="ptrA of Aspergillus oryzae"
                     /product="pyrithiamine resistance protein (Kubodera et al.,
                     2000)"
                     /label=ptrA
                     /note="useful as a dominant selectable marker in 
                     Aspergillus"
                     /translation="MSPPAAIYEPTVAATGLKGKVVVSETVPVEGASQTKLLDHFGGKW
                     DEFKFAPIRESQVSRAMTRRYFEDLDKYAESDVVIVGAGSCGLSTAYVLAKARPDLKIA
                     IVEASVSPGGGAWLGGQLFSAMVMRRPAEVFLNELGVPYEEDANPNYVVVKHASLFTST
                     LMSKVLSFPNVKLFNATAVEDLITRPTENGNPQIAGVVVNWTLVTLHHDDHSCMDPNTI
                     NAPVIISTTGHDGPFGAFCAKRLVSMGSVDKLGGMRGLDMSSAEDAIVKNTREVTKGLI
                     IGGMELSEIDGFNRMGPTFGAMVLSGVKAAEEALKVFDERQRECAE"
     CDS             4925..4936
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             join(5267..5335,5336..6127)
                     /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(5485..5504)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6298..6886
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6787..6806
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7040..7057
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    7174..7195
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(7210..7215,7216..7233,7234..7240)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7248..7264
                     /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     7253..7275
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     7272..7288
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7272..7288
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     intron          8409..8524
                     /gene="gpdA"
                     /label=gpdA intron
                     /note="intron from the Aspergillus nidulans 
                     glyceraldehyde-3-phosphate dehydrogenase gene (Punt et al.,
                     1990)"
     CDS             8545..8565
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8596..12699
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             8596..12699
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             12712..12732
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
ORIGIN
        1 tactgcagtc ggtcaagagg taactctaag tcggccattc ctttttggga ggcgctaacc
       61 ataaacggca tggtcgactt agagttagct cagggaattt agggagttat ctgcgaccac
      121 tggactctca ggctttccat tggggtacta acatagccat caaatgccgg cccgtgttta
      181 atctcctcga gatggctggg ctcacatgcg ttctactcaa aaactcttat acctaggccc
      241 tctcccaatc gtgtatgttt ggaccttcct ctctcacact ttggataaca tcaagcatat
      301 tgcagatttt gacctagtta acaaggcaca tcatgctatc acttccaaca ctcttgctga
      361 gcgcgttctg ccttgccttt ctggttctac gaagtatcca gtcccgcctg caacgccgca
      421 gaaacgccaa aaggtggggt tgtcaaccgc cagctagagc tccatcggga ttctttggca
      481 ttccgccgtt cctcggtggt cgcagataac tccctaaatt ccctgagcta actctaagtc
      541 gaccatgccg tttatggtta gcgcctccca aaaaggaatg gccgacttag agttacctct
      601 tgaccgactt tttctttcct cccccttaca tttcgttacc acaacacatt cctatatcca
      661 aactccaggt acataactag tcgaaatctc ttttaaatct agtcaagaac tagatactta
      721 accttcatac tgcttagaga tagcctttga agctcttatt tcgctctttg tctcactctc
      781 accttcctcc tcctcctcca atcctttttg ctcgggacta gtccaagact agtcaacgat
      841 tagtatgccg agcattcgcg ataacgatct ccgaaagtcc ccagagtact gtcactatct
      901 cgaggcagtt aaggacgggg agcttacgct gccggatttc aagatagtaa gccgacccga
      961 ctagctttaa gcctagttac tgactagtcc acaggacgac aatggcgtgc ctgatatcca
     1021 tccatatgaa gtctactgcc gagtgaaggg atgcctcaag cgtacagtga gtctactgct
     1081 atactagttt ctgactagtt attaactagt tgacaggttc cctctgccaa cagaaacata
     1141 ttggtcaagc acttgaagga caagaactcc cacggcatgg agtttacatt gcacaatgga
     1201 cctcccacta tgaaggaact gatggaggcc aaaggcaagt cctatctaga ttacttggtg
     1261 actagtccct gactagtcaa gtagcatggt atgaaggctt gtttgaaggc actgttctcc
     1321 caaccccgac tcctaccaag aagcgcaagc gagctgcgta agtttctgtg agtctaacta
     1381 gtgtattagc taatatatag cagcaccaag tccaaggacc acaatactaa gggtgtcgag
     1441 aattcgtgag tttcttctcc catttcaact agtccttgac tagtcactaa ctactttgca
     1501 gcaacgaggg tgaagctgga aatgatcagg acaatggcga gggcccgtaa gtacagccat
     1561 tcaatgcaga ctagttgcta actaatctgt gactagttca agtggtccgt acgccgtgca
     1621 tacccctgtg actggtagga atttgagcaa gcctgtcttg ccgcgcgatg agaaaggaaa
     1681 ggcaagttac attcagcccc gtacctagga tcagtgctaa tttataacct ctagccactc
     1741 tttatgcaga tccgccgtga gggtagcaag gcagctaaat cagctggtga gaaaggaacc
     1801 ataccctgca agacctgtcg caacgcaaag ggcaaaggta agctatccaa gctagtttgg
     1861 gactagattc taactagtct cagcaccgtg tggttcaaag ccatattgcg agttttggcg
     1921 ctttttctca tcgattgacg aggcaaaggg agcgagtatg caacctcgta agtcagactc
     1981 agacaaagac caactagtta ttgaccagtc ttctagaagg ctctgttgtg gatcttgagg
     2041 ccctggagag ttcctccaac aatccggaga caagcaagtc ctcgtcggac tagtcactaa
     2101 ctagactcta actagttgca gacatggata atgcaaaaga gacaagcaat gaagaaagtg
     2161 gtaagacatt tctcctttgt ggttctggac tagtctttga ctagtcacag tcttaaacaa
     2221 ggaaaatgag catgaaaatg aggaggaaaa ggctgctgag cccgaggaag tgcagggtga
     2281 tggcagacat ggtaggttaa taccttgtta gttattgcta gtcactgact agtcaataac
     2341 tagtctctga acaccttgca atcactccgt ttgcgcagct gaacagtggt gaggataata
     2401 gtagtaagtt attctagctt cagagttata ggagactaga tactaactag tattagttgc
     2461 aactaacctg gatctcagag actttggcct caatctagaa tctatctagt tgtcaactag
     2521 actgtggtat cattgtcttt tattttccta gtcctggaac tagcttctaa ctagtctccc
     2581 taatatgtgg ctgtcttgtt tttttttttt gtttccctac ccggatatct agtccccttc
     2641 taggttctgt taacctctcg ggctctgatt tagtttaacg caaacctgag attagtttct
     2701 aactagtctc taggttttct atccaccttt aattgtaata ataaatacaa gcaacgttta
     2761 tacgtcaaaa gcatttataa acttttaccc taaagtagct tgcttgtgtg tttagtttat
     2821 aattagtctc ttattaattt gatgtaggta agcccgccac aaatatatat ttttaacaag
     2881 ataccgtgga aaaacttcgt gctatcacaa aacagtatac aaaaaataag ctggctttcc
     2941 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     3001 cgaccccaaa aaacttgatt tgggtgatgg ttcacgtagt gggccatcgc cctgatagac
     3061 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     3121 tggaacaaca ctcaactcta tctcgggcta ttcttttgat ttataaggga ttttgccgat
     3181 ttcggtctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa
     3241 aatggggtga cgatgagccg ctcttgcatc tttgtttgta ttatactgtc tttcttgtta
     3301 cacataatta ttctagaatg ccccaccgtt acatacggga cacagccatt tacatatgca
     3361 tgtggattac gagctaacga gttcattcaa atctcagaac tatcacataa tcatcattcc
     3421 cctatcgtca aagaccgtaa gacaaatccg gttcatgcac tgaacccatt cgggtagtga
     3481 gtcatttact cagcacactc gcgctgacgc tcgtcgaaca ccttcaatgc ctcctcggca
     3541 gccttgacac cactgagaac catggcaccg aaggtagggc ccatgcggtt aaagccatca
     3601 atttcagaca gctccatacc gccgattatc aagcccttag taacctcgcg ggtgttcttg
     3661 acgatggcat cctcggccga gctcatgtcg agaccacgca tgccacctag cttgtcgacg
     3721 ctgcccatgg acaccaagcg cttcgcacag aaggcgccga atggcccatc gtgaccagtg
     3781 gtactgatga tgacaggagc gttgatagtg ttggggtcca tgcaggagtg atcatcgtgg
     3841 tgaagggtga ccagcgtcca gttgacgaca acaccagcaa tctgggggtt gccgttctcg
     3901 gtcggacggg tgatcaagtc ctcaacagcg gtagcattga agagcttgac attggggaag
     3961 gagagaacct tcgacatgag tgtcgaggta aacagggagg cgtgcttgac gacaacgtag
     4021 ttggggtttg cgtcctcttc gtaaggaaca cccagctcgt tcaggaagac ttccgcggga
     4081 cggcgcatga ccatagcaga aaagagttgg ccacccaacc aggcaccgcc acctattgca
     4141 cgttagttcc ggaaagctga gtgcaaggca atccatcatg gactactgac caggagagac
     4201 gctggcctcg acgatagcaa tcttcaggtc cggacgagcc ttggccaaga cgtacgcagt
     4261 gctcagaccg caggaaccag caccaacaat gacaacgtca ctttcagcgt acttgtccag
     4321 gtcctcaaag taacgtctgg tcatggcacg agagacctgg ctttcgcgga taggggcgaa
     4381 cttgaactcg tcccacttgc caccgaaatg gtccaacagc ttggtctgag aagctccctc
     4441 aacggggacg gtctcagaaa ccacgacctt acccttgagg ccggtagcgg ccacagtggg
     4501 ttcgtagatg gcagctggag gagacatgtt tcaagttgca atgactatca tctgttagcc
     4561 attccatcaa caggaagaac gagagaaggc atgacccttt tcgctggtat tatccagatc
     4621 aagttttagc cgtataatct cagaacgaac ccagtccatc gatgccatgt ccttctagac
     4681 taggatccta gagtctaggg cccagcttag ggagggcatg tgaatgcatc gatgactggg
     4741 aacgaacacc ggcccacgcc aaagacgtta cctaagatac cttgatcatt gtgagagtcc
     4801 agccaaaagt attccatgac ttccatcgta tgccctctag agggctaatc gaggagtgta
     4861 tttacattgt cggttggttt gggaactata gaagatggtc agttattcca atcaccaaag
     4921 gtttatcgaa gggaggaaga cttgttcagt ttcgtccgag gacttttgga attcaaatct
     4981 gagatagaga attgtgtggg atgagaggaa aaggaagaag atggtggggt tcagaaggag
     5041 gggttgagtt aaatagcatg ggttgagtca acgtgataag ggcactatac cgtatagatc
     5101 agcggcaccc gattctatcc gttccttttg ctcctcttta gctttgaccg gtgagccgga
     5161 caagaaacaa gtgaaatcat cctgacatcg gcggacgatc tcctagcttt tacatttcgt
     5221 taccaatggg atcccgtaat caattgccca ttgaaaaagg aagagtatga gtattcaaca
     5281 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
     5341 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
     5401 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
     5461 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
     5521 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
     5581 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
     5641 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     5701 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     5761 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     5821 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     5881 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     5941 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     6001 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     6061 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     6121 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     6181 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     6241 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     6301 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     6361 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     6421 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     6481 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     6541 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     6601 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     6661 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     6721 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     6781 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     6841 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     6901 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     6961 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     7021 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gcggaagagc gcccaatacg
     7081 caaaccgcct ctccccgcgc gttggccgat tcattaatgc agctggcacg acaggtttcc
     7141 cgactggaaa gcgggcagtg agcgcaacgc aattaatgtg agttagctca ctcattaggc
     7201 accccaggct ttacacttta tgcttccggc tcgtatgttg tgtggaattg tgagcggata
     7261 acaatttcac acaggaaaca gctatgacca tgattacgcc aagcttcgag taccatttaa
     7321 ttctatttgt gtttgatcga gacctaatac agcccctaca acgaccatca aagtcgtata
     7381 gctaccagtg aggaagtgga ctcaaatcga cttcagcaac atctcctgga taaactttaa
     7441 gcctaaacta tacagaataa gatggtggag agcttatacc gagctcccaa atctgtccag
     7501 atcatggttg accggtgcct ggatcttcct atagaatcat ccttattcgt tgacctagct
     7561 gattctggag tgacccagag ggtcatgact tgagcctaaa atccgccgcc tccaccattt
     7621 gtagaaaaat gtgacgaact cgtgagctct gtacagtgac cggtgactct ttctggcatg
     7681 cggagagacg gacggacgca gagagaaggg ctgagtaata agcgccactg cgccagacag
     7741 ctctggcggc tctgaggtgc agtggatgat tattaatccg ggaccggccg cccctccgcc
     7801 ccgaagtgga aaggctggtg tgcccctcgt tgaccaagaa tctattgcat catcggagaa
     7861 tatggagctt catcgaatca ccggcagtaa gcgaaggaga atgtgaagcc aggggtgtat
     7921 agccgtcggc gaaatagcat gccattaacc taggtacaga agtccaattg cttccgatct
     7981 ggtaaaagat tcacgagata gtaccttctc cgaagtaggt agagcgagta cccggcgcgt
     8041 aagctcccta attggcccat ccggcatctg tagggcgtcc aaatatcgtg cctctcctgc
     8101 tttgcccggt gtatgaaacc ggaaaggccg ctcaggagct ggccagcggc gcagaccggg
     8161 aacacaagct ggcagtcgac ccatccggtg ctctgcactc gacctgctga ggtccctcag
     8221 tccctggtag gcagctttgc cccgtctgtc cgcccggtgt gtcggcgggg ttgacaaggt
     8281 cgttgcgtca gtccaacatt tgttgccata ttttcctgct ctccccacca gctgctcttt
     8341 tcttttctct ttcttttccc atcttcagta tattcatctt cccatccaag aacctttatt
     8401 tcccctaagt aagtactttg ctacatccat actccatcct tcccatccct tattcctttg
     8461 aacctttcag ttcgagcttt cccacttcat cgcagcttga ctaacagcta ccccgcttga
     8521 gcaggcggcc gcatgatcat ggccccaaag aagaagcgga aggtcggtat ccacggagtc
     8581 ccagcagccg gccgcatgga caagaagtac tccatcggtc tcgacatcgg caccaactcc
     8641 gtcggttggg ccgtcatcac cgacgagtac aaggtcccct ccaagaagtt caaggtcctc
     8701 ggtaacaccg accgtcactc catcaagaag aacctcatcg gtgccctcct cttcgactcc
     8761 ggtgagaccg ccgaggccac ccgtctcaag cgtaccgccc gtcgtcgtta cacccgtcgt
     8821 aagaaccgta tctgctacct ccaggagatc ttctccaacg agatggctaa ggtcgatgac
     8881 tccttcttcc accgtctcga ggagtccttc ctcgtcgagg aggataagaa gcacgagcgt
     8941 caccccatct tcggtaacat cgtcgacgag gtcgcctacc acgagaagta ccccaccatc
     9001 taccacctcc gtaagaagct ggtcgactct actgataagg ctgaccttcg tctcatctac
     9061 ctcgccctcg cccacatgat caagttccgt ggtcacttcc tcatcgaggg tgacctcaac
     9121 cctgacaact ccgatgtcga caagctcttc atccagctcg ttcagactta caaccagctc
     9181 ttcgaagaga accccatcaa cgcttccggt gttgacgcca aggccatcct ctccgctcgt
     9241 ctctccaagt cccgtcgtct cgagaacctc atcgctcagc tccctggtga gaagaagaac
     9301 ggtctcttcg gtaaccttat cgccctctct ctcggtctca cccccaactt caagtccaac
     9361 ttcgacctcg ccgaggatgc caaactccag ctctccaagg acacctacga tgacgatctc
     9421 gacaacctcc tcgcccagat cggtgaccag tacgctgacc tcttccttgc cgctaagaac
     9481 ctctccgacg ccatcctcct ctccgatatc ctccgtgtca acaccgagat caccaaggct
     9541 cccctctccg cttctatgat caagcgttac gatgagcacc accaggacct cactcttctc
     9601 aaggcccttg tccgtcagca gctccctgag aagtacaagg agatcttctt cgatcagtcc
     9661 aagaacggtt acgccggtta catcgacggc ggtgcttccc aggaggagtt ctacaagttc
     9721 atcaagccca tcctcgagaa gatggacggc accgaggagc tcctcgtcaa gctcaaccgt
     9781 gaggatctcc tccgtaagca gcgtactttc gacaacggtt ccatccccca ccagatccac
     9841 cttggcgaac tccacgctat cctccgtcgt caggaggact tctacccctt cctcaaggac
     9901 aaccgtgaga agatcgagaa gatcctcacc ttccgtatcc cctactacgt tggccccctc
     9961 gcccgtggta actcccgttt cgcctggatg actcgtaagt ccgaagagac catcactccc
    10021 tggaacttcg aggaagtcgt cgataagggt gcctctgccc agtccttcat cgagcgtatg
    10081 actaacttcg acaagaacct ccctaacgag aaggtccttc ctaagcactc tctcctttac
    10141 gagtacttca ctgtttacaa cgagctcacc aaggtcaagt acgtcaccga aggtatgcgt
    10201 aagcccgcct tcctctctgg tgagcagaag aaggctatcg tcgacctcct cttcaagacc
    10261 aaccgtaagg ttaccgtcaa gcagctcaag gaagactact tcaagaagat cgagtgcttc
    10321 gactccgttg agatctccgg tgtcgaggat cgtttcaacg cttccctcgg cacctaccac
    10381 gatctcctca agatcatcaa ggacaaggac ttcctcgaca acgaggagaa cgaggacatc
    10441 cttgaggaca tcgtcctcac ccttaccctc ttcgaggatc gtgagatgat cgaagaacgt
    10501 ctcaagactt acgctcacct cttcgacgac aaggtcatga agcagctcaa gcgtcgtcgt
    10561 tacactggtt ggggccgtct ctcccgtaag ctgatcaacg gcatccgtga caagcagtct
    10621 ggtaagacta tcctcgactt ccttaagtcc gatggtttcg ccaaccgtaa cttcatgcag
    10681 ctcatccacg acgactctct caccttcaag gaggacatcc agaaggctca ggtttccggt
    10741 cagggtgact cccttcacga gcacatcgct aacctcgccg gttcccccgc tatcaagaag
    10801 ggtatcctcc agaccgttaa ggtcgtcgat gagctcgtca aggttatggg tcgtcacaag
    10861 cccgagaaca tcgttatcga gatggcccgt gagaaccaga ctacccagaa gggtcagaag
    10921 aactctcgtg aacgtatgaa gcgtatcgag gagggtatca aggaactcgg ctcccagatc
    10981 cttaaggagc accccgtcga gaacacccag cttcagaacg agaagctcta cctctactac
    11041 ctccagaacg gccgtgacat gtacgttgat caggagctcg acatcaaccg tctctccgac
    11101 tacgacgtcg atcacatcgt cccccagtcc ttcctcaagg atgactctat cgacaacaag
    11161 gtcctcaccc gttccgataa gaaccgtggc aagtctgaca acgtcccctc cgaagaggtt
    11221 gtcaagaaga tgaagaacta ctggcgtcag cttctcaacg ccaagctcat cacccagcgt
    11281 aagttcgata acctcactaa ggctgaacgt ggtggcctct ctgagctcga caaggccggc
    11341 ttcatcaagc gtcagcttgt tgagactcgt cagatcacca agcacgtcgc ccagattctc
    11401 gactctcgca tgaacaccaa gtacgatgag aacgacaagc tcatccgtga ggtcaaggtt
    11461 atcactctca agtctaagct cgtctccgac ttccgtaagg acttccagtt ctacaaggtc
    11521 cgtgagatca acaactacca ccacgcccac gatgcctacc tcaacgctgt cgttggcact
    11581 gctcttatca agaagtaccc caagctggag tccgagttcg tctacggtga ctacaaggtc
    11641 tacgatgttc gtaagatgat cgctaagtcc gagcaggaga tcggtaaggc caccgctaag
    11701 tacttcttct actccaacat catgaacttc ttcaagaccg agatcaccct cgccaacggt
    11761 gagatccgta agcgtcccct catcgagacc aacggtgaga ctggtgagat cgtctgggac
    11821 aagggtcgtg acttcgccac tgtccgtaag gtcctctcca tgccccaggt caacatcgtc
    11881 aagaagaccg aggtccagac cggtggcttc tccaaggaat ctatcctccc caagcgtaac
    11941 tccgacaagc tcatcgctcg caagaaggac tgggacccca agaagtacgg cggtttcgac
    12001 tctcccactg tcgcttactc cgtcctcgtt gtcgccaagg tcgagaaggg taagtctaag
    12061 aagctcaagt ccgtcaagga gctcctcggc atcaccatca tggagcgttc ctccttcgag
    12121 aagaacccca tcgacttcct cgaggccaag ggttacaagg aggtcaagaa ggacctcatc
    12181 atcaagctgc ccaagtactc cctcttcgag cttgagaacg gtcgtaagcg tatgctcgct
    12241 tccgccggtg agctccagaa gggtaacgag ctcgctctcc cctccaagta cgtcaacttc
    12301 ctctacctcg cctcccacta cgagaagctc aagggttctc ccgaagacaa cgagcagaag
    12361 cagctcttcg tcgagcagca caagcactac cttgatgaga tcatcgagca gatctccgag
    12421 ttctccaagc gtgtcatcct cgccgacgct aacctcgata aggtcctctc cgcttacaac
    12481 aagcaccgtg acaagcccat ccgtgagcag gctgagaaca tcatccacct cttcactctc
    12541 accaaccttg gtgcccctgc tgccttcaag tacttcgaca ccaccatcga ccgtaagcgt
    12601 tacacctcta ccaaggaggt cctcgacgcc actctcatcc accagtccat taccggtctc
    12661 tacgagactc gtatcgacct ctctcagctc ggtggtgact cccgtgctga ccccaagaag
    12721 aagcgtaagg tctgagctag gggaattccg gaccggatcc acttaacgtt actgaaatca
    12781 tcaaacagct tgacgaatct ggatataaga tcgttggtgt cgatgtcagc tccggagttg
    12841 agacaaatgg tgttcaggat ctcgataaga tacgttcatt tgtccaagca gcaaagagtg
    12901 ccttctagtg atttaatagc tccatgtcaa caagaataaa acgcgtttcg ggtttacctc
    12961 ttccagatac agctcatctg caatgcatta atgcattgac tgcaacctag taacggtacc
    13021 a
//
