LOCUS       sequence-449561-       11982 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Disruption of S. cerevisiae type LEU2 genes.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11982)
  AUTHORS   Kiyokawa K, Sakuma T, Moriguchi K, Sugiyama M, Akao T, Yamamoto T, 
            Suzuki K
  TITLE     Conversion of polyploid and alloploid Saccharomyces sensu stricto 
            strains to leu2 mutants by genome DNA editing.
  JOURNAL   Appl Microbiol Biotechnol. 2024 Jul 12;108(1):416. doi: 
            10.1007/s00253-024-13242-y.
  PUBMED    38995331
REFERENCE   2  (bases 1 to 11982)
  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: "Appl 
            Microbiol Biotechnol."; date: "2024-07-12"; volume: "108(1):416. 
            doi"; pages: " 10.1007/s00253-024-13242-y"
COMMENT     Sequence Label: pYAMTrGCsgScLEU2
FEATURES             Location/Qualifiers
     source          1..11982
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    10..513
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     555..573
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(611..633)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     733..752
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     gene            808..2164
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        808..1151
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1152..1961
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1219..1238)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      1967..2164
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     rep_origin      complement(2246..2701)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2383..2404)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2595..2614
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2827..2849
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2841..2858
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2842..2858
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2868..2887
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2868..2886
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     2919..2935
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2919..2935
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    3009..3030
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3045..3050,3051..3068,3069..3075)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3083..3099
                     /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     3107..3123
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        3144..3162
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        3193..3461
                     /label=SNR52 promoter
                     /note="promoter for the S. cerevisiae small nucleolar RNA 
                     gene SNR52"
     misc_RNA        3482..3556
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     terminator      3561..3580
                     /label=SUP4 terminator
                     /note="transcription terminator for the S. cerevisiae SUP4 
                     tRNA gene"
     terminator      complement(3698..3885)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     3806..3830
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(3913..3933)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(6328..8046)
                     /codon_start=1
                     /product="N-terminal portion of Streptococcus pyogenes Cas9
                     (Zetsche et al., 2015)"
                     /label=Cas9(N)
                     /translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIE"
     regulatory      8043..8052
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(8081..8747)
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     complement(8188..8211)
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     primer_bind     complement(8792..8808)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8793..8809)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(8837..8855)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(8876..8892)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    8900..8916
                     /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(8924..8930,8931..8948,8949..8954))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     misc_feature    8992..9016
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             complement(9283..10077)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(9394..9413)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     10004..10023
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     rep_origin      10491..11262
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     CDS             11263..11925
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
ORIGIN
        1 gtccttttca tcacgtgcta taaaaataat tataatttaa attttttaat ataaatatat
       61 aaattaaaaa tagaaagtaa aaaaagaaat taaagaaaaa atagtttttg ttttccgaag
      121 atgtaaaaga ctctaggggg atcgccaaca aatactacct tttatcttgc tcttcctgct
      181 ctcaggtatt aatgccgaat tgtttcatct tgtctgtgta gaagaccaca cacgaaaatc
      241 ctgtgatttt acattttact tatcgttaat cgaatgtata tctatttaat ctgcttttct
      301 tgtctaataa atatatatgt aaagtacgct ttttgttgaa attttttaaa cctttgttta
      361 tttttttttc ttcattccgt aactcttcta ccttctttat ttactttcta aaatccaaat
      421 acaaaacata aaaataaata aacacagagt aaattcccaa attattccat cattaaaaga
      481 tacgaggcgc gtgtaagtta caggcaagcg atccgtccta agaaaccatt attatcatga
      541 cattaaccta taaaaatagg cgtatcacga ggccctttcg tctcgcgcgt ttcggtgatg
      601 acggtgaaaa cctctgacac atgcagctcc cggagacggt cacagcttgt ctgtaagcgg
      661 atgccgggag cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg tgtcggggct
      721 ggcttaacta tgcggcatca gagcagattg tactgagagt gcaccatagt ttagcttgcc
      781 tcgtccccgc cgggtcaccc ggccagcgac atggaggccc agaataccct ccttgacagt
      841 cttgacgtgc gcagctcagg ggcatgatgt gactgtcgcc cgtacattta gcccatacat
      901 ccccatgtat aatcatttgc atccatacat tttgatggcc gcacggcgcg aagcaaaaat
      961 tacggctcct cgctgcagac ctgcgagcag ggaaacgctc ccctcacaga cgcgttgaat
     1021 tgtccccacg ccgcgcccct gtagagaaat ataaaaggtt aggatttgcc actgaggttc
     1081 ttctttcata tacttccttt taaaatcttg ctaggataca gttctcacat cacatccgaa
     1141 cataaacaac catgggtaag gaaaagactc acgtttcgag gccgcgatta aattccaaca
     1201 tggatgctga tttatatggg tataaatggg ctcgcgataa tgtcgggcaa tcaggtgcga
     1261 caatctatcg attgtatggg aagcccgatg cgccagagtt gtttctgaaa catggcaaag
     1321 gtagcgttgc caatgatgtt acagatgaga tggtcagact aaactggctg acggaattta
     1381 tgcctcttcc gaccatcaag cattttatcc gtactcctga tgatgcatgg ttactcacca
     1441 ctgcgatccc cggcaaaaca gcattccagg tattagaaga atatcctgat tcaggtgaaa
     1501 atattgttga tgcgctggca gtgttcctgc gccggttgca ttcgattcct gtttgtaatt
     1561 gtccttttaa cagcgatcgc gtatttcgtc tcgctcaggc gcaatcacga atgaataacg
     1621 gtttggttga tgcgagtgat tttgatgacg agcgtaatgg ctggcctgtt gaacaagtct
     1681 ggaaagaaat gcataagctt ttgccattct caccggattc agtcgtcact catggtgatt
     1741 tctcacttga taaccttatt tttgacgagg ggaaattaat aggttgtatt gatgttggac
     1801 gagtcggaat cgcagaccga taccaggatc ttgccatcct atggaactgc ctcggtgagt
     1861 tttctccttc attacagaaa cggctttttc aaaaatatgg tattgataat cctgatatga
     1921 ataaattgca gtttcatttg atgctcgatg agtttttcta atcagtactg acaataaaaa
     1981 gattcttgtt ttcaagaact tgtcatttgt atagtttttt tatattgtag ttgttctatt
     2041 ttaatcaaat gttagcgtga tttatatttt ttttcgcctc gacatcatct gcccagatgc
     2101 gaagttaagt gcgcagaaag taatatcatg cgtcaatcgt atgtgaatgc tggtcgctat
     2161 actgctgtcg attcgatact aacgccgcca tccagtgtcg aaaacaatac cgcacagatg
     2221 cgtaaggaga aaataccgca tcaggaaatt gtaagcgtta atattttgtt aaaattcgcg
     2281 ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg caaaatccct
     2341 tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg gaacaagagt
     2401 ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat
     2461 ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg ccgtaaagca
     2521 ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa gccggcgaac
     2581 gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta
     2641 gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct acagggcgcg
     2701 tccattcgcc attcaggctg cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc
     2761 tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg gtaacgccag
     2821 ggttttccca gtcacgacgt tgtaaaacga cggccagtga gcgcgcgtaa tacgactcac
     2881 tatagggcga attggagctc caccgcggtg gcggccgctc tagaactagt ggatcccccg
     2941 ggctgcagga attcgatctg gcacgacagg tttcccgact ggaaagcggg cagtgagcgc
     3001 aacgcaatta atgtgagtta gctcactcat taggcacccc aggctttaca ctttatgctt
     3061 ccggctcgta tgttgtgtgg aattgtgagc ggataacaat ttcacacagg aaacagctat
     3121 gaccatgatt acgccaagcg cgcaattaac cctcactaaa gggaacaaaa gctggagctc
     3181 caccgcggac aatctttgaa aagataatgt atgattatgc tttcactcat atttatacag
     3241 aaacttgatg ttttctttcg agtatataca aggtgattac atgtacgttt gaagtacaac
     3301 tctagatttt gtagtgccct cttgggctag cggtaaaggt gcgcattttt tcacacccta
     3361 caatgttctg ttcaaaagat tttggtcaaa cgctgtagaa gtgaaagttg gtgcgcatgt
     3421 ttcggcgttc gaaacttctc cgcagtgaaa gataaatgat caaggaccaa ataggcaatg
     3481 ggttttagag ctagaaatag caagttaaaa taaggctagt ccgttatcaa cttgaaaaag
     3541 tggcaccgag tcggtggtgc tttttttgtt ttttatgtct tcgagtcatg taattagtta
     3601 taagcatgtg agctaagaca ctgtaattgc caatctaaac gataccacgg ccgctctaga
     3661 gaaatgggga gcgatttgca ggcatttgct cggcatgccg gtagaggtgt ggtcaataag
     3721 agcgacctca tgctatacct gagaaagcaa cctgacctac aggaaagagt tactcaagaa
     3781 taagaatttt cgttttaaaa cctaagagtc actttaaaat ttgtatacac ttattttttt
     3841 tataacttat ttaataataa aaatcataaa tcataagaaa ttcgcctcga gagtctagga
     3901 tccggaacta cctaccttgc gctttttctt gggagctccc tcatcccctc cgagctgtga
     3961 gaggtcaatc cgagtttcgt aaagtccggt gatgctctgg tggatgaggg tggcgtccag
     4021 cacttctttg gtggaggtat agcgtttccg gtcgatggtg gtgtcgaagt acttgaatgc
     4081 ggcaggggca ccaagattgg tgagggtgaa caggtggatg atattctcag cctgctcccg
     4141 aattggtttg tctctgtgct tattgtatgc gctaaggact ttatcgaggt tggcatcagc
     4201 gagaatgacg cgtttagaaa actcggaaat ctgctcaatg atctcgtcca gataatgctt
     4261 atgctgctcc acaaacagtt gcttttgctc gttgtcctcg gggctcccct tcagcttctc
     4321 gtaatggctt gcaaggtaga ggaaattcac gtacttggag gggagagcca gctcattacc
     4381 cttctgaagt tcgccagcgg aggcgagcat ccgcttccgc ccattctcca gttcgaacag
     4441 gctatacttt gggagcttga tgataaggtc cttcttgact tccttgtaac ctttggcttc
     4501 gagaaagtca attgggttct tctcgaagga gctgcgctcc ataatggtga tacccagcag
     4561 ttccttcacg ctcttgagct tcttagactt tcctttttcc accttagcca cgaccaggac
     4621 agaataagcc acagttggtg aatcgaatcc gccgtacttc ttagggtccc aatccttctt
     4681 ccttgcaatg agcttgtcgg agttgcgctt ggggaggatt gattccttag aaaagcctcc
     4741 ggtttgcacc tcggttttct tcacgatgtt cacttgaggc atagaaagca ctttgcgcac
     4801 ggtagcgaaa tctctgccct tgtcccacac gatttcgcca gtctccccat tagtctcgat
     4861 gagcggcctc ttccggatct caccgtttgc aagggtgatt tcggtcttga agaaattcat
     4921 gatgttgctg tagaagaagt actttgcggt ggcctttccg atctcctgct cagatttggc
     4981 aatcatcttg cgcacgtcgt agaccttgta gtctccgtac acgaactcgc tttccagctt
     5041 tgggtacttc ttgattaggg cggttcccac cacggcattc agatatgcgt catgagcatg
     5101 atgatagtta ttgatctccc gcactttgta gaattgaaag tcctttctga aatcgctgac
     5161 cagcttggac ttcagggtaa tcaccttcac ctcgcggata agtttgtcgt tttcgtcgta
     5221 cttggtattc atgcgtgagt caaggatttg ggctacatgt ttagtaatct gccgagtctc
     5281 cacaagttgc cgtttaatga atcctgcttt gtccagttcg ctcagtccgc ccctctcagc
     5341 cttagtgaga ttgtcgaact tccgctgagt gattagcttt gcgttcagaa gctggcgcca
     5401 atagttcttc atctttttca cgacttcctc tgagggcacg ttatctgact ttccccggtt
     5461 cttgtctgag cgagtcagca ccttattgtc aattgagtcg tctttgagga agctctgagg
     5521 gacgatgtgg tccacgtcgt agtctgagag cctgttgata tcaagctctt ggtccacata
     5581 catatcgcgt ccattttgaa ggtagtacag atagagcttc tcattctgaa gctgggtgtt
     5641 ttccaccggg tgctctttaa ggatctgtga cccgagctcc ttaatccctt cctcaattct
     5701 tttcatccgc tccctgctgt tcttctgtcc cttctgggta gtctggtttt ccctggccat
     5761 ctcgatgaca atgttctctg gtttatggcg acccatcacc ttgaccagct cgtccacgac
     5821 tttcacggtc tggagaatac ccttcttaat agcgggtgaa ccagccagat ttgcgatatg
     5881 ctcgtgcagt gagtcacctt gtccagacac ttgggccttc tggatgtcct ccttaaaggt
     5941 cagggaatca tcatggataa gttgcatgaa gttgcgatta gcgaatccgt cgctcttaag
     6001 gaaatcaaga atggtctttc cgctctgttt gtcccggatt ccgttgatga gtttgcggct
     6061 aaggcgtccc catccggtat atcttctccg tttaagttgc ttcatcactt tatcgtcgaa
     6121 cagatgggcg taggtcttaa gcctctcctc gatcatttcc cggtcctcga acagagtgag
     6181 agtcaggaca atgtcctcaa ggatgtcctc tttctcctca ttgtccagaa aatccttgtc
     6241 cttgatgatc ttaagcagat catggtaggt acccaggctt gcgttgaacc gatcttcgac
     6301 cccgctaatc tccacagagt cgaagcattc aattttcttg aaatagtcct ccttcagctg
     6361 cttcacggtc acctttctat tggtcttgaa cagaaggtcc acaattgcct tcttttgttc
     6421 gcctgacagg aatgcgggct tcctcatccc ttcggtgaca tatttcacct tggtcagctc
     6481 gttgtacacg gtaaagtact cgtagagcag ggaatgtttc ggcaggacct tctcattggg
     6541 gaggttctta tcgaagttag tcatccgttc gatgaatgac tgagcgctgg ctcccttgtc
     6601 caccacctct tcgaagttcc agggagtgat agtttcttct gactttctgg tcatccaagc
     6661 aaaccggcta tttcctctgg cgaggggtcc cacgtagtag gggatgcgga aagtgagaat
     6721 tttctcaatc ttctccctgt tgtccttaag aaaggggtag aagtcctctt ggcgccgaag
     6781 gatggcgtga agctccccaa gatggatctg gtggggaatg ctcccattgt cgaaagtgcg
     6841 ctgcttccgc agcagatcct cccgattcag tttcacgagg agctcttcgg tgccgtccat
     6901 cttctcaagg ataggtttga taaacttata aaattcctct tgggaggcac ctccgtcgat
     6961 gtagccggcg tatccattct ttgactggtc aaagaagatt tccttgtact tctccggaag
     7021 ctgctgccgg acgagtgcct taagcagggt caggtcctgg tggtgctcat cataccgctt
     7081 aatcattgaa gctgaaagag gggccttggt gatttcagtg ttcacgcgga gaatgtcaga
     7141 cagcaggatg gcgtcagaaa gattcttagc agcaaggaaa aggtcggcgt attggtcacc
     7201 gatctgggcc aggaggttgt cgagatcgtc atcataggtg tccttggaca gttgcagctt
     7261 ggcatcctct gccaggtcga aattagactt gaaattggga gtcagcccaa gtgaaagagc
     7321 aatcaggttc ccgaacagtc catttttctt ttcacccggc agctgagcga taagattctc
     7381 aagtctgcgg ctctttgaca gccgggctga cagaatggcc ttggcatcga ctccgcttgc
     7441 attgatgggg ttctcttcaa agagctggtt gtaagtctgc accagctgga tgaacagttt
     7501 atccacatcg ctattgtcag gattgaggtc gccctcgatc aggaagtggc ctctgaactt
     7561 aatcatgtgg gccagggcca ggtagataag ccggaggtct gccttgtcgg ttgagtccac
     7621 cagctttttg cgaagatggt agatggttgg gtatttctcg tgataggcca cctcgtcgac
     7681 gatattaccg aagatgggat gccgttcatg cttcttatcc tcttccacga ggaatgattc
     7741 ttccagccta tggaagaagc tgtcgtcgac ctttgccatt tcgttggaga atatctcttg
     7801 aaggtaacaa atccggttct tcctcctggt gtaccgccgt ctagcagtcc gcttgaggcg
     7861 ggttgcttca gcggtctccc ctgaatcaaa gaggagagct ccgatcagat ttttcttgat
     7921 ggaatgccgg tcggtgttcc cgagcacctt gaacttctta gagggcacct tgtactcgtc
     7981 agtgatgacg gcccacccga cgctattagt cccgatgtcc agtccgatag aatacttctt
     8041 gtccatggtg gcgagatcta attcgatcca tatcagatct tttgtttgtt tatgtgtgtt
     8101 tattcgaaac taagttcttg gtgttttaaa actaaaaaaa agactaacta taaaagtaga
     8161 atttaagaag tttaagaaat agatttacag aattacaatc aatacctacc gtctttatat
     8221 acttattagt caagtagggg aataatttca gggaactggt ttcaaccttt tttttcagct
     8281 ttttccaaat cagagagagc agaaggtaat agaaggtgta agaaaatgag atagatacat
     8341 gcgtgggtca attgccttgt gtcatcattt actccaggca ggttgcatca ctccattgag
     8401 gttgtgcccg ttttttgcct gtttgtgccc ctgttctctg tagttgcgct aagagaatgg
     8461 acctatgaac tgatggttgg tgaagaaaac aatattttgg tgctgggatt cttttttttt
     8521 ctggatgcca gcttaaaaag cgggctccat tatatttagt ggatgccagg aataaactgt
     8581 tcacccagac acctacgatg ttatatattc tgtgtaaccc gccccctatt ttgggcatgt
     8641 acgggttaca gcagaattaa aaggctaatt ttttgactaa ataaagttag gaaaatcact
     8701 actattaatt atttacgtat tctttgaaat ggcagtattg ataatgataa actcgaactg
     8761 actagtagac tgaattcgat atcaagctta tcgataccgt cgacctcgag ggggggcccg
     8821 gtaccagctt ttgttccctt tagtgagggt taatttcgag cttggcgtaa tcatggtcat
     8881 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     8941 gcataaagtg taaagcctgg ggtgcctaat gagtgaggta cggatcgatc ctgacaggat
     9001 atattggcgg gtaaacctaa gagaaaagag cgtttattag aataatcgga tatttaaaag
     9061 ggcgtgaaaa ggtttatccg ttcgtccatt tgtatgtgca tgccaaccac agggttcccc
     9121 agatctggcg ccggccagcg agacgagcaa gattggccgc ggtcacactg cttccggtag
     9181 tcaataaacc ggtaaaccag caatagacat aagcggctat ttaacgaccc tgccctgaac
     9241 cgacgaccgg gtcgaatttg ctttcgaacc ccagagtccc gctcagaaga actcgtcaag
     9301 aaggcgatag aaggcgatgc gctgcgaatc gggagcggcg ataccgtaaa gcacgaggaa
     9361 gcggtcagcc cattcgccgc caagctcttc agcaatatca cgggtagcca acgctatgtc
     9421 ctgatagcgg tccgccacac ccagccggcc acagtcgatg aatccagaaa agcggccatt
     9481 ttccaccatg atattcggca agcaggcatc gccatgggtc acgacgagat cctcgccgtc
     9541 gggcatgcgc gccttgagcc tggcgaacag ttcggctggc gcgagcccct gatgctcttc
     9601 gtccagatca tcctgatcga caagaccggc ttccatccga gtacgtgctc gctcgatgcg
     9661 atgtttcgct tggtggtcga atgggcaggt agccggatca agcgtatgca gccgccgcat
     9721 tgcatcagcc atgatggata ctttctcggc aggagcaagg tgagatgaca ggagatcctg
     9781 ccccggcact tcgcccaata gcagccagtc ccttcccgct tcagtgacaa cgtcgagcac
     9841 agctgcgcaa ggaacgcccg tcgtggccag ccacgatagc cgcgctgcct cgtcctgcag
     9901 ttcattcagg gcaccggaca ggtcggtctt gacaaaaaga accgggcgcc cctgcgctga
     9961 cagccggaac acggcggcat cagagcagcc gattgtctgt tgtgcccagt catagccgaa
    10021 tagcctctcc acccaagcgg ccggagaacc tgcgtgcaat ccatcttgtt caatcatgcg
    10081 aaacgatcct catcctgtct cttgatcaga tcttgatccc ctgcgccatc agatccttgg
    10141 cggcaagaaa gccatccagt ttactttgca gggcttccca accttaccag agggcgcccc
    10201 agctggcaat tccggttcgc ttgctgtcca taaaaccgcc cagtctagct atcgccatgt
    10261 aagcccactg cctcgagact agttggtgtc caaccggctc gacgggggca gcgcaaggcg
    10321 gtgcctccgg cgggccactc aatgcttgag tatactcact agactttgct tcgcaaagtc
    10381 gtgaccgcct acggcggctg cggcgcccta cgggcttgct ctccgggctt cgccctgcgc
    10441 ggtcgctgcg ctcccttgcc agcccgtgga tatgtggacg atggccgcga gcggccaccg
    10501 gctggctcgc ttcgctcggc ccgtggacaa ccctgctgga caagctgatg gacaggctgc
    10561 gcctgcccac gagcttgacc acagggattg cccaccggct acccagcctt cgaccacata
    10621 cccaccggct ccaactgcgc ggcctgcggc cttgccccat caattttttt aattttctct
    10681 ggggaaaagc ctccggcctg cggcctgcgc gcttcgcttg ccggttggac accaagtgga
    10741 aggcgggtca aggctcgcgc agcgaccgcg cagcggcttg gccttgacgc gcctggaacg
    10801 acccaagcct atgcgagtgg gggcagtcga agggcgaagc ccgcccgcct gccccccgag
    10861 cctcacggcg gcgagtgcgg gggttccaag ggggcagcgc caccttgggc aaggccgaag
    10921 gccgcgcagt cgatcaacaa gccccggagg ggccactttt tgccggaggg ggagccgcgc
    10981 cgaaggcgtg ggggaacccc gcaggggtgc ccttctttgg gcaccaaaga actagatata
    11041 gggcgaaatg cgaaagactt aaaaatcaac aacttaaaaa aggggggtac gcaacagctc
    11101 attgcggcac cccccgcaat agctcattgc gtaggttaaa gaaaatctgt aattgactgc
    11161 cacttttacg caacgcataa ttgttgtcgc gctgccgaaa agttgcagct gattgcgcat
    11221 ggtgccgcaa ccgtgcggca cccctaccgc atggagataa gcatggccac gcagtccaga
    11281 gaaatcggca ttcaagccaa gaacaagccc ggtcactggg tgcaaacgga acgcaaagcg
    11341 catgaggcgt gggccgggct tattgcgagg aaacccacgg cggcaatgct gctgcatcac
    11401 ctcgtggcgc agatgggcca ccagaacgcc gtggtggtca gccagaagac actttccaag
    11461 ctcatcggac gttctttgcg gacggtccaa tacgcagtca aggacttggt ggccgagcgc
    11521 tggatctccg tcgtgaagct caacggcccc ggcaccgtgt cggcctacgt ggtcaatgac
    11581 cgcgtggcgt ggggccagcc ccgcgaccag ttgcgcctgt cggtgttcag tgccgccgtg
    11641 gtggttgatc acgacgacca ggacgaatcg ctgttggggc atggcgacct gcgccgcatc
    11701 ccgaccctgt atccgggcga gcagcaacta ccgaccggcc ccggcgagga gccgcccagc
    11761 cagcccggca ttccgggcat ggaaccagac ctgccagcct tgaccgaaac ggaggaatgg
    11821 gaacggcgcg ggcagcagcg cctgccgatg cccgatgagc cgtgttttct ggacgatggc
    11881 gagccgttgg agccgccgac acgggtcacg ctgccgcgcc ggtagcactt gggttgcgca
    11941 gcaacccgta agtgcgctgt tccagactat cggctgtagc cg
//
