LOCUS       sequence-446582-       12843 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Disruption of S. cerevisiae type LEU2 gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12843)
  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 12843)
  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: pYAMTr2GCsgScLEU2
FEATURES             Location/Qualifiers
     source          1..12843
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      29..1375
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     primer_bind     1416..1434
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1472..1494)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     1594..1613
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     gene            1669..3025
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        1669..2012
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             2013..2822
                     /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(2080..2099)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      2828..3025
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     rep_origin      complement(3107..3562)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3244..3265)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3456..3475
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3688..3710
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3702..3719
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3703..3719
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3729..3748
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3729..3747
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     3780..3796
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3780..3796
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    3870..3891
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3906..3911,3912..3929,3930..3936)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3944..3960
                     /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     3968..3984
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        4005..4023
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        4054..4322
                     /label=SNR52 promoter
                     /note="promoter for the S. cerevisiae small nucleolar RNA 
                     gene SNR52"
     misc_RNA        4343..4417
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     terminator      4422..4441
                     /label=SUP4 terminator
                     /note="transcription terminator for the S. cerevisiae SUP4 
                     tRNA gene"
     terminator      complement(4559..4746)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     4667..4691
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(4774..4794)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(7189..8907)
                     /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      8904..8913
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(8942..9608)
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     complement(9049..9072)
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     primer_bind     complement(9653..9669)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9654..9670)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     promoter        complement(9698..9716)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(9737..9753)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    9761..9777
                     /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(9785..9791,9792..9809,9810..9815))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     misc_feature    9853..9877
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             complement(10144..10938)
                     /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(10255..10274)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     10865..10884
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     rep_origin      11352..12123
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     CDS             12124..12786
                     /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 tcacgtgagt gccacctgaa cgaagcatct gtgcttcatt ttgtagaaca
       61 aaaatgcaac gcgagagcgc taatttttca aacaaagaat ctgagctgca tttttacaga
      121 acagaaatgc aacgcgaaag cgctatttta ccaacgaaga atctgtgctt catttttgta
      181 aaacaaaaat gcaacgcgag agcgctaatt tttcaaacaa agaatctgag ctgcattttt
      241 acagaacaga aatgcaacgc gagagcgcta ttttaccaac aaagaatcta tacttctttt
      301 ttgttctaca aaaatgcatc ccgagagcgc tatttttcta acaaagcatc ttagattact
      361 ttttttctcc tttgtgcgct ctataatgca gtctcttgat aactttttgc actgtaggtc
      421 cgttaaggtt agaagaaggc tactttggtg tctattttct cttccataaa aaaagcctga
      481 ctccacttcc cgcgtttact gattactagc gaagctgcgg gtgcattttt tcaagataaa
      541 ggcatccccg attatattct ataccgatgt ggattgcgca tactttgtga acagaaagtg
      601 atagcgttga tgattcttca ttggtcagaa aattatgaac ggtttcttct attttgtctc
      661 tatatactac gtataggaaa tgtttacatt ttcgtattgt tttcgattca ctctatgaat
      721 agttcttact acaatttttt tgtctaaaga gtaatactag agataaacat aaaaaatgta
      781 gaggtcgagt ttagatgcaa gttcaaggag cgaaaggtgg atgggtaggt tatataggga
      841 tatagcacag agatatatag caaagagata cttttgagca atgtttgtgg aagcggtatt
      901 cgcaatattt tagtagctcg ttacagtccg gtgcgttttt ggttttttga aagtgcgtct
      961 tcagagcgct tttggttttc aaaagcgctc tgaagttcct atactttcta gctagagaat
     1021 aggaacttcg gaataggaac ttcaaagcgt ttccgaaaac gagcgcttcc gaaaatgcaa
     1081 cgcgagctgc gcacatacag ctcactgttc acgtcgcacc tatatctgcg tgttgcctgt
     1141 atatatatat acatgagaag aacggcatag tgcgtgttta tgcttaaatg cgtacttata
     1201 tgcgtctatt tatgtaggat gaaaggtagt ctagtacctc ctgtgatatt atcccattcc
     1261 atgcggggta tcgtatgctt ccttcagcac taccctttag ctgttctata tgctgccact
     1321 cctcaattgg attagtctca tccttcaatg ctatcatttc ctttgatatt ggatcatact
     1381 aagaaaccat tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc
     1441 gtctcgcgcg tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg
     1501 tcacagcttg tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg
     1561 gtgttggcgg gtgtcggggc tggcttaact atgcggcatc agagcagatt gtactgagag
     1621 tgcaccatag tttagcttgc ctcgtccccg ccgggtcacc cggccagcga catggaggcc
     1681 cagaataccc tccttgacag tcttgacgtg cgcagctcag gggcatgatg tgactgtcgc
     1741 ccgtacattt agcccataca tccccatgta taatcatttg catccataca ttttgatggc
     1801 cgcacggcgc gaagcaaaaa ttacggctcc tcgctgcaga cctgcgagca gggaaacgct
     1861 cccctcacag acgcgttgaa ttgtccccac gccgcgcccc tgtagagaaa tataaaaggt
     1921 taggatttgc cactgaggtt cttctttcat atacttcctt ttaaaatctt gctaggatac
     1981 agttctcaca tcacatccga acataaacaa ccatgggtaa ggaaaagact cacgtttcga
     2041 ggccgcgatt aaattccaac atggatgctg atttatatgg gtataaatgg gctcgcgata
     2101 atgtcgggca atcaggtgcg acaatctatc gattgtatgg gaagcccgat gcgccagagt
     2161 tgtttctgaa acatggcaaa ggtagcgttg ccaatgatgt tacagatgag atggtcagac
     2221 taaactggct gacggaattt atgcctcttc cgaccatcaa gcattttatc cgtactcctg
     2281 atgatgcatg gttactcacc actgcgatcc ccggcaaaac agcattccag gtattagaag
     2341 aatatcctga ttcaggtgaa aatattgttg atgcgctggc agtgttcctg cgccggttgc
     2401 attcgattcc tgtttgtaat tgtcctttta acagcgatcg cgtatttcgt ctcgctcagg
     2461 cgcaatcacg aatgaataac ggtttggttg atgcgagtga ttttgatgac gagcgtaatg
     2521 gctggcctgt tgaacaagtc tggaaagaaa tgcataagct tttgccattc tcaccggatt
     2581 cagtcgtcac tcatggtgat ttctcacttg ataaccttat ttttgacgag gggaaattaa
     2641 taggttgtat tgatgttgga cgagtcggaa tcgcagaccg ataccaggat cttgccatcc
     2701 tatggaactg cctcggtgag ttttctcctt cattacagaa acggcttttt caaaaatatg
     2761 gtattgataa tcctgatatg aataaattgc agtttcattt gatgctcgat gagtttttct
     2821 aatcagtact gacaataaaa agattcttgt tttcaagaac ttgtcatttg tatagttttt
     2881 ttatattgta gttgttctat tttaatcaaa tgttagcgtg atttatattt tttttcgcct
     2941 cgacatcatc tgcccagatg cgaagttaag tgcgcagaaa gtaatatcat gcgtcaatcg
     3001 tatgtgaatg ctggtcgcta tactgctgtc gattcgatac taacgccgcc atccagtgtc
     3061 gaaaacaata ccgcacagat gcgtaaggag aaaataccgc atcaggaaat tgtaagcgtt
     3121 aatattttgt taaaattcgc gttaaatttt tgttaaatca gctcattttt taaccaatag
     3181 gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg gttgagtgtt
     3241 gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt caaagggcga
     3301 aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc aagttttttg
     3361 gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg atttagagct
     3421 tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa aggagcgggc
     3481 gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc cgccgcgctt
     3541 aatgcgccgc tacagggcgc gtccattcgc cattcaggct gcgcaactgt tgggaagggc
     3601 gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa agggggatgt gctgcaaggc
     3661 gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg acggccagtg
     3721 agcgcgcgta atacgactca ctatagggcg aattggagct ccaccgcggt ggcggccgct
     3781 ctagaactag tggatccccc gggctgcagg aattcgatct ggcacgacag gtttcccgac
     3841 tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca ttaggcaccc
     3901 caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag cggataacaa
     3961 tttcacacag gaaacagcta tgaccatgat tacgccaagc gcgcaattaa ccctcactaa
     4021 agggaacaaa agctggagct ccaccgcgga caatctttga aaagataatg tatgattatg
     4081 ctttcactca tatttataca gaaacttgat gttttctttc gagtatatac aaggtgatta
     4141 catgtacgtt tgaagtacaa ctctagattt tgtagtgccc tcttgggcta gcggtaaagg
     4201 tgcgcatttt ttcacaccct acaatgttct gttcaaaaga ttttggtcaa acgctgtaga
     4261 agtgaaagtt ggtgcgcatg tttcggcgtt cgaaacttct ccgcagtgaa agataaatga
     4321 tcaaggacca aataggcaat gggttttaga gctagaaata gcaagttaaa ataaggctag
     4381 tccgttatca acttgaaaaa gtggcaccga gtcggtggtg ctttttttgt tttttatgtc
     4441 ttcgagtcat gtaattagtt ataagcatgt gagctaagac actgtaattg ccaatctaaa
     4501 cgataccacg gccgctctag agaaatgggg agcgatttgc aggcatttgc tcggcatgcc
     4561 ggtagaggtg tggtcaataa gagcgacctc atgctatacc tgagaaagca acctgaccta
     4621 caggaaagag ttactcaaga ataagaattt tcgttttaaa acctaagagt cactttaaaa
     4681 tttgtataca cttatttttt ttataactta tttaataata aaaatcataa atcataagaa
     4741 attcgcctcg agagtctagg atccggaact acctaccttg cgctttttct tgggagctcc
     4801 ctcatcccct ccgagctgtg agaggtcaat ccgagtttcg taaagtccgg tgatgctctg
     4861 gtggatgagg gtggcgtcca gcacttcttt ggtggaggta tagcgtttcc ggtcgatggt
     4921 ggtgtcgaag tacttgaatg cggcaggggc accaagattg gtgagggtga acaggtggat
     4981 gatattctca gcctgctccc gaattggttt gtctctgtgc ttattgtatg cgctaaggac
     5041 tttatcgagg ttggcatcag cgagaatgac gcgtttagaa aactcggaaa tctgctcaat
     5101 gatctcgtcc agataatgct tatgctgctc cacaaacagt tgcttttgct cgttgtcctc
     5161 ggggctcccc ttcagcttct cgtaatggct tgcaaggtag aggaaattca cgtacttgga
     5221 ggggagagcc agctcattac ccttctgaag ttcgccagcg gaggcgagca tccgcttccg
     5281 cccattctcc agttcgaaca ggctatactt tgggagcttg atgataaggt ccttcttgac
     5341 ttccttgtaa cctttggctt cgagaaagtc aattgggttc ttctcgaagg agctgcgctc
     5401 cataatggtg atacccagca gttccttcac gctcttgagc ttcttagact ttcctttttc
     5461 caccttagcc acgaccagga cagaataagc cacagttggt gaatcgaatc cgccgtactt
     5521 cttagggtcc caatccttct tccttgcaat gagcttgtcg gagttgcgct tggggaggat
     5581 tgattcctta gaaaagcctc cggtttgcac ctcggttttc ttcacgatgt tcacttgagg
     5641 catagaaagc actttgcgca cggtagcgaa atctctgccc ttgtcccaca cgatttcgcc
     5701 agtctcccca ttagtctcga tgagcggcct cttccggatc tcaccgtttg caagggtgat
     5761 ttcggtcttg aagaaattca tgatgttgct gtagaagaag tactttgcgg tggcctttcc
     5821 gatctcctgc tcagatttgg caatcatctt gcgcacgtcg tagaccttgt agtctccgta
     5881 cacgaactcg ctttccagct ttgggtactt cttgattagg gcggttccca ccacggcatt
     5941 cagatatgcg tcatgagcat gatgatagtt attgatctcc cgcactttgt agaattgaaa
     6001 gtcctttctg aaatcgctga ccagcttgga cttcagggta atcaccttca cctcgcggat
     6061 aagtttgtcg ttttcgtcgt acttggtatt catgcgtgag tcaaggattt gggctacatg
     6121 tttagtaatc tgccgagtct ccacaagttg ccgtttaatg aatcctgctt tgtccagttc
     6181 gctcagtccg cccctctcag ccttagtgag attgtcgaac ttccgctgag tgattagctt
     6241 tgcgttcaga agctggcgcc aatagttctt catctttttc acgacttcct ctgagggcac
     6301 gttatctgac tttccccggt tcttgtctga gcgagtcagc accttattgt caattgagtc
     6361 gtctttgagg aagctctgag ggacgatgtg gtccacgtcg tagtctgaga gcctgttgat
     6421 atcaagctct tggtccacat acatatcgcg tccattttga aggtagtaca gatagagctt
     6481 ctcattctga agctgggtgt tttccaccgg gtgctcttta aggatctgtg acccgagctc
     6541 cttaatccct tcctcaattc ttttcatccg ctccctgctg ttcttctgtc ccttctgggt
     6601 agtctggttt tccctggcca tctcgatgac aatgttctct ggtttatggc gacccatcac
     6661 cttgaccagc tcgtccacga ctttcacggt ctggagaata cccttcttaa tagcgggtga
     6721 accagccaga tttgcgatat gctcgtgcag tgagtcacct tgtccagaca cttgggcctt
     6781 ctggatgtcc tccttaaagg tcagggaatc atcatggata agttgcatga agttgcgatt
     6841 agcgaatccg tcgctcttaa ggaaatcaag aatggtcttt ccgctctgtt tgtcccggat
     6901 tccgttgatg agtttgcggc taaggcgtcc ccatccggta tatcttctcc gtttaagttg
     6961 cttcatcact ttatcgtcga acagatgggc gtaggtctta agcctctcct cgatcatttc
     7021 ccggtcctcg aacagagtga gagtcaggac aatgtcctca aggatgtcct ctttctcctc
     7081 attgtccaga aaatccttgt ccttgatgat cttaagcaga tcatggtagg tacccaggct
     7141 tgcgttgaac cgatcttcga ccccgctaat ctccacagag tcgaagcatt caattttctt
     7201 gaaatagtcc tccttcagct gcttcacggt cacctttcta ttggtcttga acagaaggtc
     7261 cacaattgcc ttcttttgtt cgcctgacag gaatgcgggc ttcctcatcc cttcggtgac
     7321 atatttcacc ttggtcagct cgttgtacac ggtaaagtac tcgtagagca gggaatgttt
     7381 cggcaggacc ttctcattgg ggaggttctt atcgaagtta gtcatccgtt cgatgaatga
     7441 ctgagcgctg gctcccttgt ccaccacctc ttcgaagttc cagggagtga tagtttcttc
     7501 tgactttctg gtcatccaag caaaccggct atttcctctg gcgaggggtc ccacgtagta
     7561 ggggatgcgg aaagtgagaa ttttctcaat cttctccctg ttgtccttaa gaaaggggta
     7621 gaagtcctct tggcgccgaa ggatggcgtg aagctcccca agatggatct ggtggggaat
     7681 gctcccattg tcgaaagtgc gctgcttccg cagcagatcc tcccgattca gtttcacgag
     7741 gagctcttcg gtgccgtcca tcttctcaag gataggtttg ataaacttat aaaattcctc
     7801 ttgggaggca cctccgtcga tgtagccggc gtatccattc tttgactggt caaagaagat
     7861 ttccttgtac ttctccggaa gctgctgccg gacgagtgcc ttaagcaggg tcaggtcctg
     7921 gtggtgctca tcataccgct taatcattga agctgaaaga ggggccttgg tgatttcagt
     7981 gttcacgcgg agaatgtcag acagcaggat ggcgtcagaa agattcttag cagcaaggaa
     8041 aaggtcggcg tattggtcac cgatctgggc caggaggttg tcgagatcgt catcataggt
     8101 gtccttggac agttgcagct tggcatcctc tgccaggtcg aaattagact tgaaattggg
     8161 agtcagccca agtgaaagag caatcaggtt cccgaacagt ccatttttct tttcacccgg
     8221 cagctgagcg ataagattct caagtctgcg gctctttgac agccgggctg acagaatggc
     8281 cttggcatcg actccgcttg cattgatggg gttctcttca aagagctggt tgtaagtctg
     8341 caccagctgg atgaacagtt tatccacatc gctattgtca ggattgaggt cgccctcgat
     8401 caggaagtgg cctctgaact taatcatgtg ggccagggcc aggtagataa gccggaggtc
     8461 tgccttgtcg gttgagtcca ccagcttttt gcgaagatgg tagatggttg ggtatttctc
     8521 gtgataggcc acctcgtcga cgatattacc gaagatggga tgccgttcat gcttcttatc
     8581 ctcttccacg aggaatgatt cttccagcct atggaagaag ctgtcgtcga cctttgccat
     8641 ttcgttggag aatatctctt gaaggtaaca aatccggttc ttcctcctgg tgtaccgccg
     8701 tctagcagtc cgcttgaggc gggttgcttc agcggtctcc cctgaatcaa agaggagagc
     8761 tccgatcaga tttttcttga tggaatgccg gtcggtgttc ccgagcacct tgaacttctt
     8821 agagggcacc ttgtactcgt cagtgatgac ggcccacccg acgctattag tcccgatgtc
     8881 cagtccgata gaatacttct tgtccatggt ggcgagatct aattcgatcc atatcagatc
     8941 ttttgtttgt ttatgtgtgt ttattcgaaa ctaagttctt ggtgttttaa aactaaaaaa
     9001 aagactaact ataaaagtag aatttaagaa gtttaagaaa tagatttaca gaattacaat
     9061 caatacctac cgtctttata tacttattag tcaagtaggg gaataatttc agggaactgg
     9121 tttcaacctt ttttttcagc tttttccaaa tcagagagag cagaaggtaa tagaaggtgt
     9181 aagaaaatga gatagataca tgcgtgggtc aattgccttg tgtcatcatt tactccaggc
     9241 aggttgcatc actccattga ggttgtgccc gttttttgcc tgtttgtgcc cctgttctct
     9301 gtagttgcgc taagagaatg gacctatgaa ctgatggttg gtgaagaaaa caatattttg
     9361 gtgctgggat tctttttttt tctggatgcc agcttaaaaa gcgggctcca ttatatttag
     9421 tggatgccag gaataaactg ttcacccaga cacctacgat gttatatatt ctgtgtaacc
     9481 cgccccctat tttgggcatg tacgggttac agcagaatta aaaggctaat tttttgacta
     9541 aataaagtta ggaaaatcac tactattaat tatttacgta ttctttgaaa tggcagtatt
     9601 gataatgata aactcgaact gactagtaga ctgaattcga tatcaagctt atcgataccg
     9661 tcgacctcga gggggggccc ggtaccagct tttgttccct ttagtgaggg ttaatttcga
     9721 gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc
     9781 cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgaggt
     9841 acggatcgat cctgacagga tatattggcg ggtaaaccta agagaaaaga gcgtttatta
     9901 gaataatcgg atatttaaaa gggcgtgaaa aggtttatcc gttcgtccat ttgtatgtgc
     9961 atgccaacca cagggttccc cagatctggc gccggccagc gagacgagca agattggccg
    10021 cggtcacact gcttccggta gtcaataaac cggtaaacca gcaatagaca taagcggcta
    10081 tttaacgacc ctgccctgaa ccgacgaccg ggtcgaattt gctttcgaac cccagagtcc
    10141 cgctcagaag aactcgtcaa gaaggcgata gaaggcgatg cgctgcgaat cgggagcggc
    10201 gataccgtaa agcacgagga agcggtcagc ccattcgccg ccaagctctt cagcaatatc
    10261 acgggtagcc aacgctatgt cctgatagcg gtccgccaca cccagccggc cacagtcgat
    10321 gaatccagaa aagcggccat tttccaccat gatattcggc aagcaggcat cgccatgggt
    10381 cacgacgaga tcctcgccgt cgggcatgcg cgccttgagc ctggcgaaca gttcggctgg
    10441 cgcgagcccc tgatgctctt cgtccagatc atcctgatcg acaagaccgg cttccatccg
    10501 agtacgtgct cgctcgatgc gatgtttcgc ttggtggtcg aatgggcagg tagccggatc
    10561 aagcgtatgc agccgccgca ttgcatcagc catgatggat actttctcgg caggagcaag
    10621 gtgagatgac aggagatcct gccccggcac ttcgcccaat agcagccagt cccttcccgc
    10681 ttcagtgaca acgtcgagca cagctgcgca aggaacgccc gtcgtggcca gccacgatag
    10741 ccgcgctgcc tcgtcctgca gttcattcag ggcaccggac aggtcggtct tgacaaaaag
    10801 aaccgggcgc ccctgcgctg acagccggaa cacggcggca tcagagcagc cgattgtctg
    10861 ttgtgcccag tcatagccga atagcctctc cacccaagcg gccggagaac ctgcgtgcaa
    10921 tccatcttgt tcaatcatgc gaaacgatcc tcatcctgtc tcttgatcag atcttgatcc
    10981 cctgcgccat cagatccttg gcggcaagaa agccatccag tttactttgc agggcttccc
    11041 aaccttacca gagggcgccc cagctggcaa ttccggttcg cttgctgtcc ataaaaccgc
    11101 ccagtctagc tatcgccatg taagcccact gcctcgagac tagttggtgt ccaaccggct
    11161 cgacgggggc agcgcaaggc ggtgcctccg gcgggccact caatgcttga gtatactcac
    11221 tagactttgc ttcgcaaagt cgtgaccgcc tacggcggct gcggcgccct acgggcttgc
    11281 tctccgggct tcgccctgcg cggtcgctgc gctcccttgc cagcccgtgg atatgtggac
    11341 gatggccgcg agcggccacc ggctggctcg cttcgctcgg cccgtggaca accctgctgg
    11401 acaagctgat ggacaggctg cgcctgccca cgagcttgac cacagggatt gcccaccggc
    11461 tacccagcct tcgaccacat acccaccggc tccaactgcg cggcctgcgg ccttgcccca
    11521 tcaatttttt taattttctc tggggaaaag cctccggcct gcggcctgcg cgcttcgctt
    11581 gccggttgga caccaagtgg aaggcgggtc aaggctcgcg cagcgaccgc gcagcggctt
    11641 ggccttgacg cgcctggaac gacccaagcc tatgcgagtg ggggcagtcg aagggcgaag
    11701 cccgcccgcc tgccccccga gcctcacggc ggcgagtgcg ggggttccaa gggggcagcg
    11761 ccaccttggg caaggccgaa ggccgcgcag tcgatcaaca agccccggag gggccacttt
    11821 ttgccggagg gggagccgcg ccgaaggcgt gggggaaccc cgcaggggtg cccttctttg
    11881 ggcaccaaag aactagatat agggcgaaat gcgaaagact taaaaatcaa caacttaaaa
    11941 aaggggggta cgcaacagct cattgcggca ccccccgcaa tagctcattg cgtaggttaa
    12001 agaaaatctg taattgactg ccacttttac gcaacgcata attgttgtcg cgctgccgaa
    12061 aagttgcagc tgattgcgca tggtgccgca accgtgcggc acccctaccg catggagata
    12121 agcatggcca cgcagtccag agaaatcggc attcaagcca agaacaagcc cggtcactgg
    12181 gtgcaaacgg aacgcaaagc gcatgaggcg tgggccgggc ttattgcgag gaaacccacg
    12241 gcggcaatgc tgctgcatca cctcgtggcg cagatgggcc accagaacgc cgtggtggtc
    12301 agccagaaga cactttccaa gctcatcgga cgttctttgc ggacggtcca atacgcagtc
    12361 aaggacttgg tggccgagcg ctggatctcc gtcgtgaagc tcaacggccc cggcaccgtg
    12421 tcggcctacg tggtcaatga ccgcgtggcg tggggccagc cccgcgacca gttgcgcctg
    12481 tcggtgttca gtgccgccgt ggtggttgat cacgacgacc aggacgaatc gctgttgggg
    12541 catggcgacc tgcgccgcat cccgaccctg tatccgggcg agcagcaact accgaccggc
    12601 cccggcgagg agccgcccag ccagcccggc attccgggca tggaaccaga cctgccagcc
    12661 ttgaccgaaa cggaggaatg ggaacggcgc gggcagcagc gcctgccgat gcccgatgag
    12721 ccgtgttttc tggacgatgg cgagccgttg gagccgccga cacgggtcac gctgccgcgc
    12781 cggtagcact tgggttgcgc agcaacccgt aagtgcgctg ttccagacta tcggctgtag
    12841 ccg
//
