LOCUS       sequence-146695-        9838 bp    DNA     circular SYN 22-AUG-2025
DEFINITION  Expresses Cas9 protein, under the control of the TEF1 promoter, and 
            CYC terminator. KanMX resistance.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9838)
  AUTHORS   Jessop-Fabre MM, Jakociunas T, Stovicek V, Dai Z, Jensen MK, 
            Keasling JD, Borodina I
  TITLE     EasyClone-MarkerFree: A vector toolkit for marker-less integration 
            of genes into Saccharomyces cerevisiae via CRISPR-Cas9.
  JOURNAL   Biotechnol J. 2016 Aug;11(8):1110-7. doi: 10.1002/biot.201600147. 
            Epub 2016 Jun 23.
  PUBMED    27166612
REFERENCE   2  (bases 1 to 9838)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 22, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1002/biot.201600147"; journalName: "Biotechnol J"; date: 
            "2016-08"; volume: "11"; issue: "8"; pages: "1110-7"
COMMENT     Sequence Label: pCfB2312 (TEF1p-Cas9-CYC1t_kanMX)
FEATURES             Location/Qualifiers
     source          1..9838
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    59..562
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        599..703
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(704..772,773..1564)
                     /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(922..941)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1735..2323
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2224..2243
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2477..2494
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        join(2647..2652,2653..2670,2671..2677)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2685..2701
                     /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     2690..2712
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2709..2725
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2709..2725
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     2744..2764
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        2746..2764
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        2782..3180
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     primer_bind     3184..3200
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3184..3200
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     CDS             3212..7315
                     /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             3212..7315
                     /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             3212..7315
                     /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             7328..7348
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     terminator      7364..7611
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(7380..7398)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(7629..7648)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(7630..7648)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(7658..7675)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(7658..7674)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7667..7689)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      7819..8199
                     /direction=RIGHT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7906..7925)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     8116..8137
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     protein_bind    complement(8251..8284)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     gene            complement(8347..9703)
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     terminator      complement(8347..8544)
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(8550..9359)
                     /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     9273..9292
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(9360..9703)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     protein_bind    complement(9758..9791)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
ORIGIN
        1 ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc ttaggacgga
       61 tcgcttgcct gtaacttaca cgcgcctcgt atcttttaat gatggaataa tttgggaatt
      121 tactctgtgt ttatttattt ttatgttttg tatttggatt ttagaaagta aataaagaag
      181 gtagaagagt tacggaatga agaaaaaaaa ataaacaaag gtttaaaaaa tttcaacaaa
      241 aagcgtactt tacatatata tttattagac aagaaaagca gattaaatag atatacattc
      301 gattaacgat aagtaaaatg taaaatcaca ggattttcgt gtgtggtctt ctacacagac
      361 aagatgaaac aattcggcat taatacctga gagcaggaag agcaagataa aaggtagtat
      421 ttgttggcga tccccctaga gtcttttaca tcttcggaaa acaaaaacta ttttttcttt
      481 aatttctttt tttactttct atttttaatt tatatattta tattaaaaaa tttaaattat
      541 aattattttt atagcacgtg atgaaaagga cccaggtggc acttttcggg gaaatgtgcg
      601 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
      661 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
      721 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
      781 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
      841 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
      901 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
      961 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     1021 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     1081 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     1141 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     1201 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     1261 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     1321 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     1381 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     1441 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     1501 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     1561 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     1621 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     1681 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     1741 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     1801 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     1861 agcgcagata ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa
     1921 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     1981 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     2041 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     2101 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     2161 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     2221 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     2281 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     2341 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     2401 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     2461 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa
     2521 accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga
     2581 ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tacctcactc attaggcacc
     2641 ccaggcttta cactttatgc ttccggctcc tatgttgtgt ggaattgtga gcggataaca
     2701 atttcacaca ggaaacagct atgaccatga ttacgccaag cgcgcaatta accctcacta
     2761 aagggaacaa aagctggagc tcatagcttc aaaatgtttc tactcctttt ttactcttcc
     2821 agattttctc ggactccgcg catcgccgta ccacttcaaa acacccaagc acagcatact
     2881 aaatttcccc tctttcttcc tctagggtgt cgttaattac ccgtactaaa ggtttggaaa
     2941 agaaaaaaga gaccgcctcg tttctttttc ttcgtcgaaa aaggcaataa aaatttttat
     3001 cacgtttctt tttcttgaaa attttttttt tgattttttt ctctttcgat gacctcccat
     3061 tgatatttaa gttaataaac ggtcttcaat ttctcaagtt tcagtttcat ttttcttgtt
     3121 ctattacaac tttttttact tcttgctcat tagaaagaaa gcatagcaat ctaatctaag
     3181 ttttctagaa ctagtggatc ccccgggaaa aatggacaag aagtactcca ttgggctcga
     3241 tatcggcaca aacagcgtcg gttgggccgt cattacggac gagtacaagg tgccgagcaa
     3301 aaaattcaaa gttctgggca ataccgatcg ccacagcata aagaagaacc tcattggcgc
     3361 cctcctgttc gactccgggg agacggccga agccacgcgg ctcaaaagaa cagcacggcg
     3421 cagatatacc cgcagaaaga atcggatctg ctacctgcag gagatcttta gtaatgagat
     3481 ggctaaggtg gatgactctt tcttccatag gctggaggag tcctttttgg tggaggagga
     3541 taaaaagcac gagcgccacc caatctttgg caatatcgtg gacgaggtgg cgtaccatga
     3601 aaagtaccca accatatatc atctgaggaa gaagcttgta gacagtactg ataaggctga
     3661 cttgcggttg atctatctcg cgctggcgca tatgatcaaa tttcggggac acttcctcat
     3721 cgagggggac ctgaacccag acaacagcga tgtcgacaaa ctctttatcc aactggttca
     3781 gacttacaat cagcttttcg aagagaaccc gatcaacgca tccggagttg acgccaaagc
     3841 aatcctgagc gctaggctgt ccaaatcccg gcggctcgaa aacctcatcg cacagctccc
     3901 tggggagaag aagaacggcc tgtttggtaa tcttatcgcc ctgtcactcg ggctgacccc
     3961 caactttaaa tctaacttcg acctggccga agatgccaag cttcaactga gcaaagacac
     4021 ctacgatgat gatctcgaca atctgctggc ccagatcggc gaccagtacg cagacctttt
     4081 tttggcggca aagaacctgt cagacgccat tctgctgagt gatattctgc gagtgaacac
     4141 ggagatcacc aaagctccgc tgagcgctag tatgatcaag cgctatgatg agcaccacca
     4201 agacttgact ttgctgaagg cccttgtcag acagcaactg cctgagaagt acaaggaaat
     4261 tttcttcgat cagtctaaaa atggctacgc cggatacatt gacggcggag caagccagga
     4321 ggaattttac aaatttatta agcccatctt ggaaaaaatg gacggcaccg aggagctgct
     4381 ggtaaagctt aacagagaag atctgttgcg caaacagcgc actttcgaca atggaagcat
     4441 cccccaccag attcacctgg gcgaactgca cgctatcctc aggcggcaag aggatttcta
     4501 cccctttttg aaagataaca gggaaaagat tgagaaaatc ctcacatttc ggatacccta
     4561 ctatgtaggc cccctcgccc ggggaaattc cagattcgcg tggatgactc gcaaatcaga
     4621 agagaccatc actccctgga acttcgagga agtcgtggat aagggggcct ctgcccagtc
     4681 cttcatcgaa aggatgacta actttgataa aaatctgcct aacgaaaagg tgcttcctaa
     4741 acactctctg ctgtacgagt acttcacagt ttataacgag ctcaccaagg tcaaatacgt
     4801 cacagaaggg atgagaaagc cagcattcct gtctggagag cagaagaaag ctatcgtgga
     4861 cctcctcttc aagacgaacc ggaaagttac cgtgaaacag ctcaaagaag actatttcaa
     4921 aaagattgaa tgtttcgact ctgttgaaat cagcggagtg gaggatcgct tcaacgcatc
     4981 cctgggaacg tatcacgatc tcctgaaaat cattaaagac aaggacttcc tggacaatga
     5041 ggagaacgag gacattcttg aggacattgt cctcaccctt acgttgtttg aagataggga
     5101 gatgattgaa gaacgcttga aaacttacgc tcatctcttc gacgacaaag tcatgaaaca
     5161 gctcaagagg cgccgatata caggatgggg gcggctgtca agaaaactga tcaatgggat
     5221 ccgagacaag cagagtggaa agacaatcct ggattttctt aagtccgatg gatttgccaa
     5281 ccggaacttc atgcagttga tccatgatga ctctctcacc tttaaggagg acatccagaa
     5341 agcacaagtt tctggccagg gggacagtct tcacgagcac atcgctaatc ttgcaggtag
     5401 cccagctatc aaaaagggaa tactgcagac cgttaaggtc gtggatgaac tcgtcaaagt
     5461 aatgggaagg cataagcccg agaatatcgt tatcgagatg gcccgagaga accaaactac
     5521 ccagaaggga cagaagaaca gtagggaaag gatgaagagg attgaagagg gtataaaaga
     5581 actggggtcc caaatcctta aggaacaccc agttgaaaac acccagcttc agaatgagaa
     5641 gctctacctg tactacctgc agaacggcag ggacatgtac gtggatcagg aactggacat
     5701 caatcggctc tccgactacg acgtggatca tatcgtgccc cagtcttttc tcaaagatga
     5761 ttctattgat aataaagtgt tgacaagatc cgataaaaat agagggaaga gtgataacgt
     5821 cccctcagaa gaagttgtca agaaaatgaa aaattattgg cggcagctgc tgaacgccaa
     5881 actgatcaca caacggaagt tcgataatct gactaaggct gaacgaggtg gcctgtctga
     5941 gttggataaa gccggcttca tcaaaaggca gcttgttgag acacgccaga tcaccaagca
     6001 cgtggcccaa attctcgatt cacgcatgaa caccaagtac gatgaaaatg acaaactgat
     6061 tcgagaggtg aaagttatta ctctgaagtc taagctggtc tcagatttca gaaaggactt
     6121 tcagttttat aaggtgagag agatcaacaa ttaccaccat gcgcatgatg cctacctgaa
     6181 tgcagtggta ggcactgcac ttatcaaaaa atatcccaag cttgaatctg aatttgttta
     6241 cggagactat aaagtgtacg atgttaggaa aatgatcgca aagtctgagc aggaaatagg
     6301 caaggccacc gctaagtact tcttttacag caatattatg aattttttca agaccgagat
     6361 tacactggcc aatggagaga ttcggaagcg accacttatc gaaacaaacg gagaaacagg
     6421 agaaatcgtg tgggacaagg gtagggattt cgcgacagtc cggaaggtcc tgtccatgcc
     6481 gcaggtgaac atcgttaaaa agaccgaagt acagaccgga ggcttctcca aggaaagtat
     6541 cctcccgaaa aggaacagcg acaagctgat cgcacgcaaa aaagattggg accccaagaa
     6601 atacggcgga ttcgattctc ctacagtcgc ttacagtgta ctggttgtgg ccaaagtgga
     6661 gaaagggaag tctaaaaaac tcaaaagcgt caaggaactg ctgggcatca caatcatgga
     6721 gcgatcaagc ttcgaaaaaa accccatcga ctttctcgag gcgaaaggat ataaagaggt
     6781 caaaaaagac ctcatcatta agcttcccaa gtactctctc tttgagcttg aaaacggccg
     6841 gaaacgaatg ctcgctagtg cgggcgagct gcagaaaggt aacgagctgg cactgccctc
     6901 taaatacgtt aatttcttgt atctggccag ccactatgaa aagctcaaag ggtctcccga
     6961 agataatgag cagaagcagc tgttcgtgga acaacacaaa cactaccttg atgagatcat
     7021 cgagcaaata agcgaattct ccaaaagagt gatcctcgcc gacgctaacc tcgataaggt
     7081 gctttctgct tacaataagc acagggataa gcccatcagg gagcaggcag aaaacattat
     7141 ccacttgttt actctgacca acttgggcgc gcctgcagcc ttcaagtact tcgacaccac
     7201 catagacaga aagcggtaca cctctacaaa ggaggtcctg gacgccacac tgattcatca
     7261 gtcaattacg gggctctatg aaacaagaat cgacctctct cagctcggtg gagacagcag
     7321 ggctgacccc aagaagaaga ggaaggtgtg atctcttctc gagtcatgta attagttatg
     7381 tcacgcttac attcacgccc tccccccaca tccgctctaa ccgaaaagga aggagttaga
     7441 caacctgaag tctaggtccc tatttatttt tttatagtta tgttagtatt aagaacgtta
     7501 tttatatttc aaatttttct tttttttctg tacagacgcg tgtacgcatg taacattata
     7561 ctgaaaacct tgcttgagaa ggttttggga cgctcgaagg ctttaatttg cggccggtac
     7621 ccaattcgcc ctatagtgag tcgtattacg cgcgctcact ggccgtcgtt ttacaacgtc
     7681 gtgactggga aaaccctggc gttacccaac ttaatcgcct tgcagcacat ccccctttcg
     7741 ccagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc
     7801 tgaatggcga atggcgcgac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     7861 ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct
     7921 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     7981 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg
     8041 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     8101 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg
     8161 tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta aattgggtgc
     8221 ataggccact agtggatctg atatcaccta ataacttcgt atagcataca ttatacgaag
     8281 ttatattaag ggttctcgag agctcgtttt cgacactgga tggcggcgtt agtatcgaat
     8341 cgacagcagt atagcgacca gcattcacat acgattgacg catgatatta ctttctgcgc
     8401 acttaacttc gcatctgggc agatgatgtc gaggcgaaaa aaaatataaa tcacgctaac
     8461 atttgattaa aatagaacaa ctacaatata aaaaaactat acaaatgaca agttcttgaa
     8521 aacaagaatc tttttattgt cagtactgat tagaaaaact catcgagcat caaatgaaac
     8581 tgcaatttat tcatatcagg attatcaata ccatattttt gaaaaagccg tttctgtaat
     8641 gaaggagaaa actcaccgag gcagttccat aggatggcaa gatcctggta tcggtctgcg
     8701 attccgactc gtccaacatc aatacaacct attaatttcc cctcgtcaaa aataaggtta
     8761 tcaagtgaga aatcaccatg agtgacgact gaatccggtg agaatggcaa aagcttatgc
     8821 atttctttcc agacttgttc aacaggccag ccattacgct cgtcatcaaa atcactcgca
     8881 tcaaccaaac cgttattcat tcgtgattgc gcctgagcga gacgaaatac gcgatcgctg
     8941 ttaaaaggac aattacaaac aggaatcgaa tgcaaccggc gcaggaacac tgccagcgca
     9001 tcaacaatat tttcacctga atcaggatat tcttctaata cctggaatgc tgttttgccg
     9061 gggatcgcag tggtgagtaa ccatgcatca tcaggagtac ggataaaatg cttgatggtc
     9121 ggaagaggca taaattccgt cagccagttt agtctgacca tctcatctgt aacatcattg
     9181 gcaacgctac ctttgccatg tttcagaaac aactctggcg catcgggctt cccatacaat
     9241 cgatagattg tcgcacctga ttgcccgaca ttatcgcgag cccatttata cccatataaa
     9301 tcagcatcca tgttggaatt taatcgcggc ctcgaaacgt gagtcttttc cttacccatg
     9361 gttgtttatg ttcggatgtg atgtgagaac tgtatcctag caagatttta aaaggaagta
     9421 tatgaaagaa gaacctcagt ggcaaatcct aaccttttat atttctctac aggggcgcgg
     9481 cgtggggaca attcaacgcg tctgtgaggg gagcgtttcc ctgctcgcag gtctgcagcg
     9541 aggagccgta atttttgctt cgcgccgtgc ggccatcaaa atgtatggat gcaaatgatt
     9601 atacatgggg atgtatgggc taaatgtacg ggcgacagtc acatcatgcc cctgagctgc
     9661 gcacgtcaag actgtcaagg agggtattct gggcctccat gtcgctggcc gggtgacccg
     9721 gcggggacga ggcaagctaa acagatctct agacctaata acttcgtata gcatacatta
     9781 tacgaagtta tattaagggt tgtcgacctg cagcgtacga agcttcagct gacgcgat
//
