LOCUS       sequence-462040-       13776 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast CEN plasmid for estradiol-inducible expression of spCas9 and 
            Ec86 reverse transcriptase..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13776)
  AUTHORS   Stuecker TN, Hood SE, Molina Pineda J, Lenaduwe S, Winter J, Sadhu 
            MJ, Lewis JA
  TITLE     Improved vectors for retron-mediated CRISPR-Cas9 genome editing in 
            Saccharomyces cerevisiae.
  JOURNAL   G3 (Bethesda). 2025 Aug 4:jkaf175. doi: 10.1093/g3journal/jkaf175.
  PUBMED    40758833
REFERENCE   2  (bases 1 to 13776)
  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: "G3 
            (Bethesda). 2025 Aug 4:jkaf175. doi: 10.1093/g3journal/jkaf175."
COMMENT     Sequence Label: pCas9-EcRT-Z3-Hyg
FEATURES             Location/Qualifiers
     source          1..13776
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    19..35
                     /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     24..46
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     43..59
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     43..59
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        80..98
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     gene            226..1801
                     /label=hphMX6
                     /note="yeast selectable marker conferring hygromycin 
                     resistance"
     promoter        226..569
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             570..1598
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     CDS             570..1598
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MGKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGR
                     GYVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDL
                     PETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHW
                     QTVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAM
                     FGDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVD
                     GNFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRA
                     KE"
     terminator      1604..1801
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     primer_bind     complement(1917..1936)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1918..1936)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1943..1960)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1943..1959)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1952..1974)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2100..2555
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2187..2206)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2397..2418
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(5887..5907)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     6583..6606
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     CDS             6649..10752
                     /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             6649..10752
                     /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             6649..10752
                     /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             10765..10785
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(10817..10835)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     terminator      10847..11036
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     11050..11072
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(11110..11128)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    11169..11672
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        11709..11813
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(11814..11882,11883..12674)
                     /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(12032..12051)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      12845..13433
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     13334..13353
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     13587..13604
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    13721..13742
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(13757..13762,13763..13776,1..4,5..11)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
ORIGIN
        1 ctcgtatgtt gtgtggaatt gtgagcggat aacaatttca cacaggaaac agctatgacc
       61 atgattacgc caagctcgaa attaaccctc actaaaggga acaaaagctg ctagagcggc
      121 cgccagctga agcttcgtac gctgcaggtc gacggatccc cgggttaatt aaggcgcgcc
      181 agatctgttt agcttgcctc gtccccgccg ggtcacccgg ccagcgacat ggaggcccag
      241 aataccctcc ttgacagtct tgacgtgcgc agctcagggg catgatgtga ctgtcgcccg
      301 tacatttagc ccatacatcc ccatgtataa tcatttgcat ccatacattt tgatggccgc
      361 acggcgcgaa gcaaaaatta cggctcctcg ctgcggacct gcgagcaggg aaacgctccc
      421 ctcacagacg cgttgaattg tccccacgcc gcgcccctgt agagaaatat aaaaggttag
      481 gatttgccac tgaggttctt ctttcatata cttcctttta aaatcttgct aggatacagt
      541 tctcacatca catccgaaca taaacaacca tgggtaaaaa gcctgaactc accgcgacgt
      601 ctgtcgagaa gtttctgatc gaaaagttcg acagcgtctc cgacctgatg cagctctcgg
      661 agggcgaaga atctcgtgct ttcagcttcg atgtaggagg gcgtggatat gtcctgcggg
      721 taaatagctg cgccgatggt ttctacaaag atcgttatgt ttatcggcac tttgcatcgg
      781 ccgcgctccc gattccggaa gtgcttgaca ttggggaatt cagcgagagc ctgacctatt
      841 gcatctcccg ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc gaactgcccg
      901 ctgttctgca gccggtcgcg gaggccatgg atgcgatcgc tgcggccgat cttagccaga
      961 cgagcgggtt cggcccattc ggaccgcaag gaatcggtca atacactaca tggcgtgatt
     1021 tcatatgcgc gattgctgat ccccatgtgt atcactggca aactgtgatg gacgacaccg
     1081 tcagtgcgtc cgtcgcgcag gctctcgatg agctgatgct ttgggccgag gactgccccg
     1141 aagtccggca cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg gacaatggcc
     1201 gcataacagc ggtcattgac tggagcgagg cgatgttcgg ggattcccaa tacgaggtcg
     1261 ccaacatctt cttctggagg ccgtggttgg cttgtatgga gcagcagacg cgctacttcg
     1321 agcggaggca tccggagctt gcaggatcgc cgcggctccg ggcgtatatg ctccgcattg
     1381 gtcttgacca actctatcag agcttggttg acggcaattt cgatgatgca gcttgggcgc
     1441 agggtcgatg cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt acacaaatcg
     1501 cccgcagaag cgcggccgtc tggaccgatg gctgtgtaga agtactcgcc gatagtggaa
     1561 accgacgccc cagcactcgt ccgagggcaa aggaataatc agtactgaca ataaaaagat
     1621 tcttgttttc aagaacttgt catttgtata gtttttttat attgtagttg ttctatttta
     1681 atcaaatgtt agcgtgattt atattttttt tcgcctcgac atcatctgcc cagatgcgaa
     1741 gttaagtgcg cagaaagtaa tatcatgcgt caatcgtatg tgaatgctgg tcgctatact
     1801 gctgtcgatt cgatactaac gccgccatcc agtgtcgaaa acgagctcga attcatcgat
     1861 gatatcagat ccactagtgg cctatgcggc cgccaccgcg gtggagctcc aattcgccct
     1921 atagtgagtc gtattacaat tcactggccg tcgttttaca acgtcgtgac tgggaaaacc
     1981 ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata
     2041 gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatgga
     2101 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     2161 tacacttgcc agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     2221 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     2281 tgctttacgg cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc
     2341 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     2401 actcttgttc caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata
     2461 agggattttg ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     2521 cgcgaatttt aacaaaatat taacgcttac aatttcctga tgcggtattt tctccttacg
     2581 catctgtgcg gtatttcaca ccgcataggg taataactga tataattaaa ttgaagctct
     2641 aatttgtgag tttagtatac atgcatgcgt cgatctcccc tcaagtggca aactgcgact
     2701 gatatgaaag taaagtatcg gaccttctat tacggaatgt atacaaacgc tacaggatac
     2761 agtgatatta aatagaaaac aagtctacaa ttaaatggca gaaagaagaa ccagcttact
     2821 actaaccacc atactcatca atacccagag catcggtgaa catctgctca aattcgaagt
     2881 cagccatatc cagcgcaccg taaggtgctg aatcatgtgg agtgaagcca ggccctggag
     2941 agtccccgtc acccaacata tctaaatcaa aatcgtctaa agcgtcagcg tgggccatgg
     3001 caacgtcttc accgtccaaa tgcagttcgc tagatgtaga ccctgctgtc gctaggtggg
     3061 attggtcggt ttcttcaacg gaggctccgc ctctagaagt tggtgcatgc aacctgtgag
     3121 cgtccagcat ttctagtaat aaatcataca gaggaacgac gtttttacac ttcatagaat
     3181 acaggtgctc catacccttg ttagacatat gtctaatgtg tgacaaaatt agtagtaact
     3241 gagctagcct ctgatgctgt tgctgcagag tcaatcctgc tttcgccata aggtgaatca
     3301 atgtatcagt gatcttgtcc aacactctat gtatatggtc tttctcttcc aggcttttca
     3361 gagttgagct taaaaatgta tagacaccgc tatttaacaa aataattgat ttcaggcaaa
     3421 cgaattcttc tccttgcaaa ttcatcattc tgaacctgga gctagtggca agaagcatat
     3481 caaaaatctc aaccatacct tctacgcatt taccttggtt cctgtccaat aataaattag
     3541 gtgcaaataa taattttaca gggtgctcca tacttctcca cactaatccg atcatcaata
     3601 tttctaacca cgcgcattca agaagatgta cctggtcatg taatgtcaag tcaacaaagc
     3661 cgggaactct tttggcccaa ttaatcatat gaactaattc cctgtctgct agatttgtca
     3721 aaagacccat catggaagcc tcggaaaaag gacgagttgg gtcgtattca ctgtacaata
     3781 taggtggttc agcatcaagt agtgcggaaa ccatttgatc agcggtcaaa gacagtgcaa
     3841 gagaattctt cttagatctt ttgatcatta atgggcttgg ccacaagttg gctgccctca
     3901 tgtctccagc gcttggatct tctagggttg aggctgctgc cggagtgccc ccacctcctg
     3961 tatggatctt tgtatgtctc tttctttcat cagatctggc gaactttctt ccacaaatgt
     4021 cacacgcgaa cggcttctcg ccggtgtggg ttctaatgtg cgtagtcaaa tggtcacttc
     4081 tgctgaagtt tctcatacag attctacatt ggaaaggctt ttggccggtg tgaatacgaa
     4141 tatgcctagt cagctcgtca gatctgctga atctcctatc acaactttct acagggcaag
     4201 cgtagggtct cgttcccatt gttaattcag taaattttcg atcttgggaa gaaaaaagca
     4261 gtaagcgtga aaaatctaaa agctgatgta gtagaagatc ctattcttta acaaagattg
     4321 accttttctt tttcttcttg gtttgagtag aaaggggaag gaagaataca agagagagga
     4381 aaaaaaggaa gataaaaaga gagcgtgata taaatgaata tatattaaac aagagagatt
     4441 gggaaggaaa ggatcaaaca aacccaaaaa tatttcaaaa aggagagaga gaggcgagtt
     4501 tggtttcaaa acggtttatt tatttatgca agaggacgtg gaagaaaaag aagaaggaag
     4561 aaaaaaattt gaaagaaaaa aacgcgtggc gggtaaagaa gaaaatggaa aatagaggcc
     4621 gggtgacaga gaaatattga gggttaattg gaaaatatgt tagggtgagg catatgtttt
     4681 taaagggatg catgcgaatt gcatatcttt ccatagattt tcaatttttg atgtctccat
     4741 ggtggtacag agaacttgta aacaattcgg tccctacatg tgaggaaatt cgctgtgaca
     4801 cttttatcac tgaactccaa atttaaaaaa tagcataaaa ttcgttatac agcaaatcta
     4861 tgtgttgcaa ttaagaacta aaagatatag agtgcatatt ttcaagaagg atagtaagct
     4921 ggcaaatcaa gtcttagcct tgttcaatgg gttcttaccg tacttctttt ccaacttaga
     4981 gatgtaggta atcaatcttc tgtgggactt agaatcaacg gacaagatga aagacaacca
     5041 acctctaaca tgttcgattt cggaggattt accgcagaaa atatggtgga tcttggctct
     5101 gatttctttg tacttttctc taccaatacc aaccttttct tgggagataa ccaaaccggt
     5161 aacttttctt tgggatcttg gaccagaaat acaggtcttt ttggagttga tgaccaaacc
     5221 ttctgatgga ataatggaga acaagaagtc tctggcctta acaactttct tcatagattg
     5281 agcagacaag gtcaagtcat cagcgtatct agtgtatatc aaacctctag aaccagcata
     5341 accttgaatt ctgtagtcca acttggagca aatcaagtta gccaattttg gagaagatgg
     5401 agcaccttgt ggcaacaagt tcttgtagca gcaaatcttg gtcaaaacag aggagatcaa
     5461 tctgttgtaa cccaaagaat ggaaaacacc gaaaaccttg ttagcagtca aagatgggaa
     5521 gaaatcttcc aagtcgatgt tcaaaatgaa gttggcacca atatgtggag tggcattgtt
     5581 caagatggat tggtgctttt caaaaccgat agaaaatggg gaggaggata acttgtccaa
     5641 gatgtttctc aaaacccaac cttgtaaagc cttcaattct ctggatggtt ggtagatagt
     5701 tctcattctc ttttctggac ccttcttttc aacagtgtag attctgtatc tgaaatcggc
     5761 ggtgtagatc aataatctca aggtttcaac agaaattctg gtagccttag acatatcgtg
     5821 caagttgttc atgactggca aacccaaatt tctcaatctg aaggtgttca agtattcagc
     5881 ggacttgacc tttcttttct tctttggtgg catttatatt gaatttatat tgaattttca
     5941 aaaattctta cttttttttt ggatggacgc aaagaagttt aataatcata ttacatggca
     6001 ttaccaccat atacatatcc atatacatat ccatatctaa tcttacttat atgttgtgga
     6061 aatgtaaaga gccccattat cttagcctaa aaaaaccttc tctttggaac tttcagtaat
     6121 acgcttaact gctcattgct atattgaagt gcggccgcgt gggcgtgcgt gggcgggcgt
     6181 gggcgtgcgt gggcgggcgt gggcgtgcgt gggcgtctag accgtgcgtc ctcgtcttca
     6241 ccggtcgcgt tcctgaaacg cagatgtgcc tcgcgccgca ctgctccgaa caataaagat
     6301 tctacaatac tagcttttat ggttatgaag aggaaaaatt ggcagtaacc tggccccaca
     6361 aaccttcaaa ttaacgaatc aaattaacaa ccataggatg ataatgcgat tagtttttta
     6421 gccttatttc tggggtaatt aatcagcgaa gcgatgattt ttgatctatt aacagatata
     6481 taaatggaaa agctgcataa ccactttaac taatactttc aacattttca gtttgtatta
     6541 cttcttattc aaatgtcata aaagtatcaa caaaaaattg ttaatatacc tctatacttt
     6601 aacgtcaagg agaaaaaact ataaactagt ggatcccccg ggaaaaaaat ggacaagaag
     6661 tactccattg ggctcgatat cggcacaaac agcgtcggtt gggccgtcat tacggacgag
     6721 tacaaggtgc cgagcaaaaa attcaaagtt ctgggcaata ccgatcgcca cagcataaag
     6781 aagaacctca ttggcgccct cctgttcgac tccggggaga cggccgaagc cacgcggctc
     6841 aaaagaacag cacggcgcag atatacccgc agaaagaatc ggatctgcta cctgcaggag
     6901 atctttagta atgagatggc taaggtggat gactctttct tccataggct ggaggagtcc
     6961 tttttggtgg aggaggataa aaagcacgag cgccacccaa tctttggcaa tatcgtggac
     7021 gaggtggcgt accatgaaaa gtacccaacc atatatcatc tgaggaagaa gcttgtagac
     7081 agtactgata aggctgactt gcggttgatc tatctcgcgc tggcgcatat gatcaaattt
     7141 cggggacact tcctcatcga gggggacctg aacccagaca acagcgatgt cgacaaactc
     7201 tttatccaac tggttcagac ttacaatcag cttttcgaag agaacccgat caacgcatcc
     7261 ggagttgacg ccaaagcaat cctgagcgct aggctgtcca aatcccggcg gctcgaaaac
     7321 ctcatcgcac agctccctgg ggagaagaag aacggcctgt ttggtaatct tatcgccctg
     7381 tcactcgggc tgacccccaa ctttaaatct aacttcgacc tggccgaaga tgccaagctt
     7441 caactgagca aagacaccta cgatgatgat ctcgacaatc tgctggccca gatcggcgac
     7501 cagtacgcag accttttttt ggcggcaaag aacctgtcag acgccattct gctgagtgat
     7561 attctgcgag tgaacacgga gatcaccaaa gctccgctga gcgctagtat gatcaagcgc
     7621 tatgatgagc accaccaaga cttgactttg ctgaaggccc ttgtcagaca gcaactgcct
     7681 gagaagtaca aggaaatttt cttcgatcag tctaaaaatg gctacgccgg atacattgac
     7741 ggcggagcaa gccaggagga attttacaaa tttattaagc ccatcttgga aaaaatggac
     7801 ggcaccgagg agctgctggt aaagcttaac agagaagatc tgttgcgcaa acagcgcact
     7861 ttcgacaatg gaagcatccc ccaccagatt cacctgggcg aactgcacgc tatcctcagg
     7921 cggcaagagg atttctaccc ctttttgaaa gataacaggg aaaagattga gaaaatcctc
     7981 acatttcgga taccctacta tgtaggcccc ctcgcccggg gaaattccag attcgcgtgg
     8041 atgactcgca aatcagaaga gaccatcact ccctggaact tcgaggaagt cgtggataag
     8101 ggggcctctg cccagtcctt catcgaaagg atgactaact ttgataaaaa tctgcctaac
     8161 gaaaaggtgc ttcctaaaca ctctctgctg tacgagtact tcacagttta taacgagctc
     8221 accaaggtca aatacgtcac agaagggatg agaaagccag cattcctgtc tggagagcag
     8281 aagaaagcta tcgtggacct cctcttcaag acgaaccgga aagttaccgt gaaacagctc
     8341 aaagaagact atttcaaaaa gattgaatgt ttcgactctg ttgaaatcag cggagtggag
     8401 gatcgcttca acgcatccct gggaacgtat cacgatctcc tgaaaatcat taaagacaag
     8461 gacttcctgg acaatgagga gaacgaggac attcttgagg acattgtcct cacccttacg
     8521 ttgtttgaag atagggagat gattgaagaa cgcttgaaaa cttacgctca tctcttcgac
     8581 gacaaagtca tgaaacagct caagaggcgc cgatatacag gatgggggcg gctgtcaaga
     8641 aaactgatca atgggatccg agacaagcag agtggaaaga caatcctgga ttttcttaag
     8701 tccgatggat ttgccaaccg gaacttcatg cagttgatcc atgatgactc tctcaccttt
     8761 aaggaggaca tccagaaagc acaagtttct ggccaggggg acagtcttca cgagcacatc
     8821 gctaatcttg caggtagccc agctatcaaa aagggaatac tgcagaccgt taaggtcgtg
     8881 gatgaactcg tcaaagtaat gggaaggcat aagcccgaga atatcgttat cgagatggcc
     8941 cgagagaacc aaactaccca gaagggacag aagaacagta gggaaaggat gaagaggatt
     9001 gaagagggta taaaagaact ggggtcccaa atccttaagg aacacccagt tgaaaacacc
     9061 cagcttcaga atgagaagct ctacctgtac tacctgcaga acggcaggga catgtacgtg
     9121 gatcaggaac tggacatcaa tcggctctcc gactacgacg tggatcatat cgtgccccag
     9181 tcttttctca aagatgattc tattgataat aaagtgttga caagatccga taaaaataga
     9241 gggaagagtg ataacgtccc ctcagaagaa gttgtcaaga aaatgaaaaa ttattggcgg
     9301 cagctgctga acgccaaact gatcacacaa cggaagttcg ataatctgac taaggctgaa
     9361 cgaggtggcc tgtctgagtt ggataaagcc ggcttcatca aaaggcagct tgttgagaca
     9421 cgccagatca ccaagcacgt ggcccaaatt ctcgattcac gcatgaacac caagtacgat
     9481 gaaaatgaca aactgattcg agaggtgaaa gttattactc tgaagtctaa gctggtctca
     9541 gatttcagaa aggactttca gttttataag gtgagagaga tcaacaatta ccaccatgcg
     9601 catgatgcct acctgaatgc agtggtaggc actgcactta tcaaaaaata tcccaagctt
     9661 gaatctgaat ttgtttacgg agactataaa gtgtacgatg ttaggaaaat gatcgcaaag
     9721 tctgagcagg aaataggcaa ggccaccgct aagtacttct tttacagcaa tattatgaat
     9781 tttttcaaga ccgagattac actggccaat ggagagattc ggaagcgacc acttatcgaa
     9841 acaaacggag aaacaggaga aatcgtgtgg gacaagggta gggatttcgc gacagtccgg
     9901 aaggtcctgt ccatgccgca ggtgaacatc gttaaaaaga ccgaagtaca gaccggaggc
     9961 ttctccaagg aaagtatcct cccgaaaagg aacagcgaca agctgatcgc acgcaaaaaa
    10021 gattgggacc ccaagaaata cggcggattc gattctccta cagtcgctta cagtgtactg
    10081 gttgtggcca aagtggagaa agggaagtct aaaaaactca aaagcgtcaa ggaactgctg
    10141 ggcatcacaa tcatggagcg atcaagcttc gaaaaaaacc ccatcgactt tctcgaggcg
    10201 aaaggatata aagaggtcaa aaaagacctc atcattaagc ttcccaagta ctctctcttt
    10261 gagcttgaaa acggccggaa acgaatgctc gctagtgcgg gcgagctgca gaaaggtaac
    10321 gagctggcac tgccctctaa atacgttaat ttcttgtatc tggccagcca ctatgaaaag
    10381 ctcaaagggt ctcccgaaga taatgagcag aagcagctgt tcgtggaaca acacaaacac
    10441 taccttgatg agatcatcga gcaaataagc gaattctcca aaagagtgat cctcgccgac
    10501 gctaacctcg ataaggtgct ttctgcttac aataagcaca gggataagcc catcagggag
    10561 caggcagaaa acattatcca cttgtttact ctgaccaact tgggcgcgcc tgcagccttc
    10621 aagtacttcg acaccaccat agacagaaag cggtacacct ctacaaagga ggtcctggac
    10681 gccacactga ttcatcagtc aattacgggg ctctatgaaa caagaatcga cctctctcag
    10741 ctcggtggag acagcagggc tgaccccaag aagaagagga aggtgtgatc tcttctcgag
    10801 tcatgtaatt agttatgtca cgcttacatt cacgccctcc ccccacatcc gctctaaccg
    10861 aaaaggaagg agttagacaa cctgaagtct aggtccctat ttattttttt atagttatgt
    10921 tagtattaag aacgttattt atatttcaaa tttttctttt ttttctgtac agacgcgtgt
    10981 acgcatgtaa cattatactg aaaaccttgc ttgagaaggt tttgggacgc tcgaaggctt
    11041 taatttcatc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga
    11101 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
    11161 cttagacgga tcgcttgcct gtaacttaca cgcgcctcgt atcttttaat gatggaataa
    11221 tttgggaatt tactctgtgt ttatttattt ttatgttttg tatttggatt ttagaaagta
    11281 aataaagaag gtagaagagt tacggaatga agaaaaaaaa ataaacaaag gtttaaaaaa
    11341 tttcaacaaa aagcgtactt tacatatata tttattagac aagaaaagca gattaaatag
    11401 atatacattc gattaacgat aagtaaaatg taaaatcaca ggattttcgt gtgtggtctt
    11461 ctacacagac aagatgaaac aattcggcat taatacctga gagcaggaag agcaagataa
    11521 aaggtagtat ttgttggcga tccccctaga gtcttttaca tcttcggaaa acaaaaacta
    11581 ttttttcttt aatttctttt tttactttct atttttaatt tatatattta tattaaaaaa
    11641 tttaaattat aattattttt atagcacgtg atgaaaagga cccaggtggc acttttcggg
    11701 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
    11761 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
    11821 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
    11881 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
    11941 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
    12001 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
    12061 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
    12121 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
    12181 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
    12241 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
    12301 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
    12361 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
    12421 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
    12481 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
    12541 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
    12601 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
    12661 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
    12721 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
    12781 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
    12841 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
    12901 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
    12961 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
    13021 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
    13081 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
    13141 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
    13201 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
    13261 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
    13321 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
    13381 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
    13441 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc
    13501 ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg
    13561 ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc
    13621 caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca
    13681 ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc
    13741 attaggcacc ccaggcttta cactttatgc ttccgg
//
