LOCUS       sequence-296525-       10002 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CRISPR/Cas9 engineering of the HO endonuclease gene with 
            nourseothricin selection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10002)
  AUTHORS   Barre B, Hallin J, Yue JX, Persson K, Mikhalev E, Irizar A, Holt S, 
            Thompson D, Molin M, Warringer J, Liti G
  TITLE     Intragenic repeat expansion in the cell wall protein gene HPF1 
            controls yeast chronological aging.
  JOURNAL   Genome Res. 2020 Apr 10. pii: gr.253351.119. doi: 
            10.1101/gr.253351.119.
  PUBMED    32277013
REFERENCE   2  (bases 1 to 10002)
  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: "Genome Res.
            2020 Apr 10. pii: gr.253351.119. doi: 10.1101/gr.253351.119."
COMMENT     Sequence Label: pL90-Nat-2xHO
FEATURES             Location/Qualifiers
     source          1..10002
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..667
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     537..560
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     ncRNA           674..710
                     /product="self-cleaving hammerhead ribozyme (Ferre-D'Amare 
                     and Scott, 2010)"
                     /label=hammerhead ribozyme
                     /ncRNA_class="hammerhead_ribozyme"
     misc_RNA        731..806
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     ncRNA           811..878
                     /product="hepatitis delta virus self-cleaving ribozyme 
                     (Ferre-D'Amare and Scott, 2010)"
                     /label=HDV ribozyme
                     /ncRNA_class="ribozyme"
     ncRNA           895..931
                     /product="self-cleaving hammerhead ribozyme (Ferre-D'Amare 
                     and Scott, 2010)"
                     /label=hammerhead ribozyme
                     /ncRNA_class="hammerhead_ribozyme"
     misc_RNA        952..1027
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     ncRNA           1032..1099
                     /product="hepatitis delta virus self-cleaving ribozyme 
                     (Ferre-D'Amare and Scott, 2010)"
                     /label=HDV ribozyme
                     /ncRNA_class="ribozyme"
     primer_bind     complement(1141..1159)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     CDS             complement(join(1593..2384,2385..2453))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGTRVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     2216..2235
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2454..2558)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2661..3249
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3150..3169
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     gene            3413..4534
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     CDS             3761..4333
                     /codon_start=1
                     /gene="Streptomyces noursei nat1"
                     /product="nourseothricin acetyltransferase"
                     /label=NrsR
                     /note="confers resistance to nourseothricin"
                     /translation="MGTTLDDTAYRYRTSVPGDAEAIEALDGSFTTDTVFRVTATGDGF
                     TLREVPVDPPLTKVFPDDESDDESDDGEDGDPDSRTFVAYGDDGDLAGFVVVSYSGWNR
                     RLTVEDIEVAPEHRGHGVGRALMGLATEFARERGAGHLWLEVTNVNAPAIHAYRRMGFT
                     LCGLDTALYDGTASDGEQALYMSMPCP"
     CDS             4912..4935
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             4939..4959
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4960..9060
                     /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
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTYKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSITHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             9064..9084
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
ORIGIN
        1 tcattatcaa tactgccatt tcaaagaata cgtaaataat taatagtagt gattttccta
       61 actttattta gtcaaaaaat tagcctttta attctgctgt aacccgtaca tgcccaaaat
      121 agggggcggg ttacacagaa tatataacat cgtaggtgtc tgggtgaaca gtttattctt
      181 ggcatccact aaatataatg gagcccgctt tttaagctgg catccagaaa aaaaaagaat
      241 cccagcacca aaatattgtt ttcttcacca accatcagtt cataggtcca ttctcttagc
      301 gcaactacag agaacagggg cacaaacagg caaaaaacgg gcacaacctc aatggagtga
      361 tgcaacctgc ctggagtaaa tgatgacaca aggcaattga cccacgcatg tatctatctc
      421 attttcttac accttctatt accttctgct ctctctgatt tggaaaaagc tgaaaaaaaa
      481 ggttgaaacc agttccctga aattattccc ctacttgact aataagtata taaagacggt
      541 aggtattgat tgtaattctg taaatctatt tcttaaactt cttaaattct acttttatag
      601 ttagtctttt ttttagtttt aaaacaccaa gaacttagtt tcgaataaac acacataaac
      661 aaacaaaaaa atactgatga gtccgtgagg acgaaacgag taagctcgtc tatttttata
      721 aagattggag gttttagagc tagaaatagc aagttaaaat aaggctagtc cgttatcaac
      781 ttgaaaaagt ggcaccgagt cggtgctttt ggccggcatg gtcccagcct cctcgctggc
      841 gccggctggg caacatgctt cggcatggcg aatgggacac agcgcaaatc tttactgatg
      901 agtccgtgag gacgaaacga gtaagctcgt ctaaagacat cgcaaacgtc agttttagag
      961 ctagaaatag caagttaaaa taaggctagt ccgttatcaa cttgaaaaag tggcaccgag
     1021 tcggtgcttt tggccggcat ggtcccagcc tcctcgctgg cgccggctgg gcaacatgct
     1081 tcggcatggc gaatgggaca caggcccctt ttcctttgtc gatatcatgt aattagttat
     1141 gtcacgctta cattcacgcc ctccccccac atccgctcta accgaaaagg aaggagttag
     1201 acaacctgaa gtctaggtcc ctatttattt ttttatagtt atgttagtat taagaacgtt
     1261 atttatattt caaatttttc ttttttttct gtacaaacgc gtgtacgcat gtaacattat
     1321 actgaaaacc ttgcttgaga aggttttggg acaagcttac gtctcacgga tcgtatatgc
     1381 cgtagcgaca atctaagaac tatgcgagga cacgctagca gctgtattca cgtagacgga
     1441 taggtatagc cagacatcag cagcatactt cgggaaccgt aggccaccga gcggccgcga
     1501 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     1561 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     1621 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     1681 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgggaccca
     1741 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     1801 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     1861 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     1921 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     1981 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     2041 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     2101 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     2161 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     2221 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     2281 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgtaccc
     2341 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     2401 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     2461 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     2521 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     2581 cctgtcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta
     2641 gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa
     2701 acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt
     2761 tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag
     2821 ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta
     2881 atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca
     2941 agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag
     3001 cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa
     3061 agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga
     3121 acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc
     3181 gggtttcccc acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc
     3241 ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt
     3301 gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtgc ggactatatg
     3361 tgaaggcatg gctatggcac ggcagacatt ccgccagatc atcaataggc acgacatgga
     3421 ggcccagaat accctccttg acagtcttga cgtgcgcagc tcaggggcat gatgtgactg
     3481 tcgcccgtac atttagccca tacatcccca tgtataatca tttgcatcca tacattttga
     3541 tggccgcacg gcgcgaagca aaaattacgg ctcctcgctg cagacctgcg agcagggaaa
     3601 cgctcccctc acagacgcgt tgaattgtcc ccacgccgcg cccctgtaga gaaatataaa
     3661 aggttaggat ttgccactga ggttcttctt tcatatactt ccttttaaaa tcttgctagg
     3721 atacagttct cacatcacat ccgaacataa acaaggatcc atgggtacta ctcttgacga
     3781 cacggcttac cggtaccgca ccagtgtccc gggggacgcc gaggccatcg aggcactgga
     3841 tgggtccttc accaccgaca ccgtcttccg cgtcaccgcc accggggacg gcttcaccct
     3901 gcgggaggtg ccggtggacc cgcccctgac caaggtgttc cccgacgacg aatcggacga
     3961 cgaatcggac gacggggagg acggcgaccc ggactcccgg acgttcgtcg cgtacgggga
     4021 cgacggcgac ctggcgggct tcgtggtcgt ctcgtactcc ggctggaacc gccggctgac
     4081 cgtcgaggac atcgaggtcg ccccggagca ccgggggcac ggggtcgggc gcgcgttgat
     4141 ggggctcgcg acggagttcg cccgcgagcg gggcgccggg cacctctggc tggaggtcac
     4201 caacgtcaac gcaccggcga tccacgcgta ccggcggatg gggttcaccc tctgcggcct
     4261 ggacaccgcc ctgtacgacg gcaccgcctc ggacggcgag caggcgctct acatgagcat
     4321 gccctgcccc taactcgagg acaataaaaa gattcttgtt ttcaagaact tgtcatttgt
     4381 atagtttttt tatattgtag ttgttctatt ttaatcaaat gttagcgtga tttatatttt
     4441 ttttcgcctc gacatcatct gcccagatgc gaagttaagt gcgcagaaag taatatcatg
     4501 cgtcaatcgt atgtgaatgc tggtcgctat actggagctc tcgagaaccc tcacctttcg
     4561 agaggacgat gcccgtgtct aaatgattcg accagcctaa gaatgttcaa caatctataa
     4621 tcagtccata gtcaacaaga gccctcaaag ttgatatcgt ttagtgatct ccctcgagat
     4681 gagcggagga tggttgcttt ttggggagcc ctaagtggta cgtgcggcga gcgggatgtt
     4741 tccccattag gcaaaggacg accgggtaac ccctcgagaa aaaaaatttt tcgccgctaa
     4801 tctggtgtta tataaagctc cccctgctcc tggaattttt ttccttgtca actcacaccg
     4861 gaaatcaagg catttcattc tgagtagttc ttaaaaacat aatcaacaat ggattataaa
     4921 gatgacgatg acaaacctcc aaaaaagaag agaaaggtcg ataagaaata ctcaataggc
     4981 ttagatatcg gcacaaatag cgtcggatgg gcggtgatca ctgatgaata taaggttccg
     5041 tctaaaaagt tcaaggttct gggaaataca gaccgccaca gtatcaaaaa aaatcttata
     5101 ggggctcttt tatttgacag tggagagaca gcggaagcga ctcgtctcaa acggacagct
     5161 cgtagaaggt atacacgtcg gaagaatcgt atttgttatc tacaggagat tttttcaaat
     5221 gagatggcga aagtagatga tagtttcttt catcgacttg aagagtcttt tttggtggaa
     5281 gaagacaaga agcatgaacg tcatcctatt tttggaaata tagtagatga agttgcttat
     5341 catgagaaat atccaactat ctatcatctg cgaaaaaaat tggtagattc tacttataaa
     5401 gcggatttgc gcttaatcta tttggcctta gcgcatatga ttaagtttcg tggtcatttt
     5461 ttgattgagg gagatttaaa tcctgataat agtgatgtgg acaaactatt tatccagttg
     5521 gtacaaacct acaatcaatt atttgaagaa aaccctatta acgcaagtgg agtagatgct
     5581 aaagcgattc tttctgcacg attgagtaaa tcaagacgat tagaaaatct cattgctcag
     5641 ctccccggtg agaagaaaaa tggcttattt gggaatctca ttgctttgtc attgggtttg
     5701 acccctaatt ttaaatcaaa ttttgatttg gcagaagatg ctaaattaca gctttcaaaa
     5761 gatacttacg atgatgattt agataattta ttggcgcaaa ttggagatca atatgctgat
     5821 ttgtttttgg cagctaagaa tttatcagat gctattttac tttcagatat cctaagagta
     5881 aatactgaaa taactaaggc tcccctatca gcttcaatga ttaaacgcta cgatgaacat
     5941 catcaagact tgactctttt aaaagcttta gttcgacaac aacttccaga aaagtataaa
     6001 gaaatctttt ttgatcaatc aaaaaacgga tatgcaggtt atattgatgg gggagctagc
     6061 caagaagaat tttataaatt tatcaaacca attttagaaa aaatggatgg tactgaggaa
     6121 ttattggtga aactaaatcg tgaagatttg ctgcgcaagc aacggacctt tgacaacggc
     6181 tctattaccc atcaaattca cttgggtgag ctgcatgcta ttttgagaag acaagaagac
     6241 ttttatccat ttttaaaaga caatcgtgag aagattgaaa aaatcttgac ttttcgaatt
     6301 ccttattatg ttggtccatt ggcgcgtggc aatagtcgtt ttgcatggat gactcggaag
     6361 tctgaagaaa caattacccc atggaatttt gaagaagttg tcgataaagg tgcttcagct
     6421 caatcattta ttgaacgcat gacaaacttt gataaaaatc ttccaaatga aaaagtacta
     6481 ccaaaacata gtttgcttta tgagtatttt acggtttata acgaattgac aaaggtcaaa
     6541 tatgttactg aaggaatgcg aaaaccagca tttctttcag gtgaacagaa gaaagccatt
     6601 gttgatttac tcttcaaaac aaatcgaaaa gtaaccgtta agcaattaaa agaagattat
     6661 ttcaaaaaaa tagaatgttt tgatagtgtt gaaatttcag gagttgaaga tagatttaat
     6721 gcttcattag gtacctacca tgatttgcta aaaattatta aagataaaga ttttttggat
     6781 aatgaagaaa atgaagatat cttagaggat attgttttaa cattgacctt atttgaagat
     6841 agggagatga ttgaggaaag acttaaaaca tatgctcacc tctttgatga taaggtgatg
     6901 aaacagctta aacgtcgccg ttatactggt tggggacgtt tgtctcgaaa attgattaat
     6961 ggtattaggg ataagcaatc tggcaaaaca atattagatt ttttgaaatc agatggtttt
     7021 gccaatcgca attttatgca gctgatccat gatgatagtt tgacatttaa agaagacatt
     7081 caaaaagcac aagtgtctgg acaaggcgat agtttacatg aacatattgc aaatttagct
     7141 ggtagccctg ctattaaaaa aggtatttta cagactgtaa aagttgttga tgaattggtc
     7201 aaagtaatgg ggcggcataa gccagaaaat atcgttattg aaatggcacg tgaaaatcag
     7261 acaactcaaa agggccagaa aaattcgcga gagcgtatga aacgaatcga agaaggtatc
     7321 aaagaattag gaagtcagat tcttaaagag catcctgttg aaaatactca attgcaaaat
     7381 gaaaagctct atctctatta tctccaaaat ggaagagaca tgtatgtgga ccaagaatta
     7441 gatattaatc gtttaagtga ttatgatgtc gatcacattg ttccacaaag tttccttaaa
     7501 gacgattcaa tagacaataa ggtcttaacg cgttctgata aaaatcgtgg taaatcggat
     7561 aacgttccaa gtgaagaagt agtcaaaaag atgaaaaact attggagaca acttctaaac
     7621 gccaagttaa tcactcaacg taagtttgat aatttaacga aagctgaacg tggaggtttg
     7681 agtgaacttg ataaagctgg ttttatcaaa cgccaattgg ttgaaactcg ccaaatcact
     7741 aagcatgtgg cacaaatttt ggatagtcgc atgaatacta aatacgatga aaatgataaa
     7801 cttattcgag aggttaaagt gattacctta aaatctaaat tagtttctga cttccgaaaa
     7861 gatttccaat tctataaagt acgtgagatt aacaattacc atcatgccca tgatgcgtat
     7921 ctaaatgccg tcgttggaac tgctttgatt aagaaatatc caaaacttga atcggagttt
     7981 gtctatggtg attataaagt ttatgatgtt cgtaaaatga ttgctaagtc tgagcaagaa
     8041 ataggcaaag caaccgcaaa atatttcttt tactctaata tcatgaactt cttcaaaaca
     8101 gaaattacac ttgcaaatgg agagattcgc aaacgccctc taatcgaaac taatggggaa
     8161 actggagaaa ttgtctggga taaagggcga gattttgcca cagtgcgcaa agtattgtcc
     8221 atgccccaag tcaatattgt caagaaaaca gaagtacaga caggcggatt ctccaaggag
     8281 tcaattttac caaaaagaaa ttcggacaag cttattgctc gtaaaaaaga ctgggatcca
     8341 aaaaaatatg gtggttttga tagtccaacg gtagcttatt cagtcctagt ggttgctaag
     8401 gtggaaaaag ggaaatcgaa gaagttaaaa tccgttaaag agttactagg gatcacaatt
     8461 atggaaagaa gttcctttga aaaaaatccg attgactttt tagaagctaa aggatataag
     8521 gaagttaaaa aagacttaat cattaaacta cctaaatata gtctttttga gttagaaaac
     8581 ggtcgtaaac ggatgctggc tagtgccgga gaattacaaa aaggaaatga gctggctctg
     8641 ccaagcaaat atgtgaattt tttatattta gctagtcatt atgaaaagtt gaagggtagt
     8701 ccagaagata acgaacaaaa acaattgttt gtggagcagc ataagcatta tttagatgag
     8761 attattgagc aaatcagtga attttctaag cgtgttattt tagcagatgc caatttagat
     8821 aaagttctta gtgcatataa caaacataga gacaaaccaa tacgtgaaca agcagaaaat
     8881 attattcatt tatttacgtt gacgaatctt ggagctcccg ctgcttttaa atattttgat
     8941 acaacaattg atcgtaaacg atatacgtct acaaaagaag ttttagatgc cactcttatc
     9001 catcaatcca tcactggtct ttatgaaaca cgcattgatt tgagtcagct aggaggtgac
     9061 cctccaaaaa agaagagaaa ggtctgagcg gttttgtata actaaataat attggaaact
     9121 aaatacgaat acccaaattt tttatctaaa ttttgccgaa agattaaaat ctgcagagat
     9181 atccgaaaca ggtaaatgga tgtttcaatc cctgtagtca gtcaggaacc catattatat
     9241 tacagtatta gtcgccgctt aggcacgcct ttaattagca aaatcaaacc ttaagtgcat
     9301 atgccgtata agggaaactc aaagaactgg catcgcaaaa atgaaaacat acgttgaaac
     9361 tacggcaaag gattggtcag atcgcttcat acagggaaag ttcggcatca acatctttgg
     9421 ataatatcag aatgagaaag aacagatacg cagtacgttt tttggtgagc tctttgcact
     9481 tctttagttc tttccatcaa tatcagtttt ttaaactttt aggactaaaa gtgatgttta
     9541 acttcaaaat gtttaaaatt ttgttcttcc cgacgttcat taagaatact aatacacttt
     9601 aataattatt ttaatatttt gttctaaata atgactttta attaaaaaag ataaaatata
     9661 aaaacatcat aataactcac cagaggttaa gaacaaaaaa acaaattaga tatctgctaa
     9721 tccaatatag ttaaatcaat ctttccttgg tataatgggt atattacata tatttcaagg
     9781 accgacactc ctaccaaata tctaaaattt accatattaa cataacatgt atataaacgt
     9841 caaatcataa tcagcactaa cgtctcacgg atcgtatatg ccgtagcgac aatctaagaa
     9901 ctatgcgagg acacgctagg gccaaggcaa aaagattcct tgattacgta agggagttag
     9961 aatcattttg aataaaaaac acgctttttc agttcgagtt ta
//
