LOCUS       sequence-150246-       11921 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Constitutive expression of Cas9 and sgRNA in Yarrowia lipolytica 
            cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11921)
  AUTHORS   Gao S, Tong Y, Wen Z, Zhu L, Ge M, Chen D, Jiang Y, Yang S
  TITLE     Multiplex gene editing of the Yarrowia lipolytica genome using the 
            CRISPR-Cas9 system.
  JOURNAL   J Ind Microbiol Biotechnol. 2016 Aug;43(8):1085-93. doi: 
            10.1007/s10295-016-1789-8. Epub 2016 Jun 27.
  PUBMED    27349768
REFERENCE   2  (bases 1 to 11921)
  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"; doi: 
            "10.1007/s10295-016-1789-8"; journalName: "J Ind Microbiol 
            Biotechnol"; date: "2016-08"; volume: "43"; issue: "8"; pages: 
            "1085-93"
COMMENT     Sequence Label: pCAS1yl
FEATURES             Location/Qualifiers
     source          1..11921
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     792..810
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     CDS             complement(859..879)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(892..4992)
                     /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
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             complement(892..4992)
                     /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
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             complement(892..4992)
                     /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
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     ncRNA           complement(6031..6098)
                     /product="hepatitis delta virus self-cleaving ribozyme 
                     (Ferre-D'Amare and Scott, 2010)"
                     /label=HDV ribozyme
                     /ncRNA_class="ribozyme"
     misc_RNA        complement(6103..6178)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     ncRNA           complement(6199..6235)
                     /product="self-cleaving hammerhead ribozyme (Ferre-D'Amare 
                     and Scott, 2010)"
                     /label=hammerhead ribozyme
                     /ncRNA_class="hammerhead_ribozyme"
     primer_bind     complement(6803..6819)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6803..6819)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6816..6838)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6827..6843
                     /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(6851..6857,6858..6875,6876..6881))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6896..6917
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(7034..7051)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7205..7793)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7285..7304)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7964..8755,8756..8824))
                     /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     8587..8606
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8825..8929)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     8997..9015
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt cgagctcggt acccgctttt
      421 cgtagataat ggaatacaaa tggatatcca gagtatacac atggatagta tacactgaca
      481 cgacaattct gtatctcttt atgttaacta ctgtgaggcg ttaaatagag cttgatatat
      541 aaaatgttac atttcacagt ctgaactttt gcagattacc taatttggta agatattaat
      601 tatgaactga aagttgatgg catccctaaa tttgatgaaa gggggatcac gcgtctgtac
      661 agaaaaaaaa gaaaaatttg aaatataaat aacgttctta atactaacat aactataaaa
      721 aaataaatag ggacctagac ttcaggttgt ctaactcctt ccttttcggt tagagcggat
      781 gtggggggag ggcgtgaatg taagcgtgac ataactaatt acatgactcg aggtccaatg
      841 actcgaggtc caaaatcaca ccttcctctt cttcttgggg tcagccctgc tgtctccacc
      901 gagctgagag aggtcgattc ttgtttcata gagccccgta attgactgat gaatcagtgt
      961 ggcgtccagg acctcctttg tagaggtgta ccgctttctg tctatggtgg tgtcgaagta
     1021 cttgaaggct gcaggcgcgc ccaagttggt cagagtaaac aagtggataa tgttttctgc
     1081 ctgctccctg atgggcttat ccctgtgctt attgtaagca gaaagcacct tatcgaggtt
     1141 agcgtcggcg aggatcactc ttttggagaa ttcgcttatt tgctcgatga tctcatcaag
     1201 gtagtgtttg tgttgttcca cgaacagctg cttctgctca ttatcttcgg gagacccttt
     1261 gagcttttca tagtggctgg ccagatacaa gaaattaacg tatttagagg gcagtgccag
     1321 ctcgttacct ttctgcagct cgcccgcact agcgagcatt cgtttccggc cgttttcaag
     1381 ctcaaagaga gagtacttgg gaagcttaat gatgaggtct tttttgacct ctttatatcc
     1441 tttcgcctcg agaaagtcga tggggttttt ttcgaagctt gatcgctcca tgattgtgat
     1501 gcccagcagt tccttgacgc ttttgagttt tttagacttc cctttctcca ctttggccac
     1561 aaccagtaca ctgtaagcga ctgtaggaga atcgaatccg ccgtatttct tggggtccca
     1621 atcttttttg cgtgcgatca gcttgtcgct gttccttttc gggaggatac tttccttgga
     1681 gaagcctccg gtctgtactt cggtcttttt aacgatgttc acctgcggca tggacaggac
     1741 cttccggact gtcgcgaaat ccctaccctt gtcccacacg atttctcctg tttctccgtt
     1801 tgtttcgata agtggtcgct tccgaatctc tccattggcc agtgtaatct cggtcttgaa
     1861 aaaattcata atattgctgt aaaagaagta cttagcggtg gccttgccta tttcctgctc
     1921 agactttgcg atcattttcc taacatcgta cactttatag tctccgtaaa caaattcaga
     1981 ttcaagcttg ggatattttt tgataagtgc agtgcctacc actgcattca ggtaggcatc
     2041 atgcgcatgg tggtaattgt tgatctctct caccttataa aactgaaagt cctttctgaa
     2101 atctgagacc agcttagact tcagagtaat aactttcacc tctcgaatca gtttgtcatt
     2161 ttcatcgtac ttggtgttca tgcgtgaatc gagaatttgg gccacgtgct tggtgatctg
     2221 gcgtgtctca acaagctgcc ttttgatgaa gccggcttta tccaactcag acaggccacc
     2281 tcgttcagcc ttagtcagat tatcgaactt ccgttgtgtg atcagtttgg cgttcagcag
     2341 ctgccgccaa taatttttca ttttcttgac aacttcttct gaggggacgt tatcactctt
     2401 ccctctattt ttatcggatc ttgtcaacac tttattatca atagaatcat ctttgagaaa
     2461 agactggggc acgatatgat ccacgtcgta gtcggagagc cgattgatgt ccagttcctg
     2521 atccacgtac atgtccctgc cgttctgcag gtagtacagg tagagcttct cattctgaag
     2581 ctgggtgttt tcaactgggt gttccttaag gatttgggac cccagttctt ttataccctc
     2641 ttcaatcctc ttcatccttt ccctactgtt cttctgtccc ttctgggtag tttggttctc
     2701 tcgggccatc tcgataacga tattctcggg cttatgcctt cccattactt tgacgagttc
     2761 atccacgacc ttaacggtct gcagtattcc ctttttgata gctgggctac ctgcaagatt
     2821 agcgatgtgc tcgtgaagac tgtccccctg gccagaaact tgtgctttct ggatgtcctc
     2881 cttaaaggtg agagagtcat catggatcaa ctgcatgaag ttccggttgg caaatccatc
     2941 ggacttaaga aaatccagga ttgtctttcc actctgcttg tctcggatcc cattgatcag
     3001 ttttcttgac agccgccccc atcctgtata tcggcgcctc ttgagctgtt tcatgacttt
     3061 gtcgtcgaag agatgagcgt aagttttcaa gcgttcttca atcatctccc tatcttcaaa
     3121 caacgtaagg gtgaggacaa tgtcctcaag aatgtcctcg ttctcctcat tgtccaggaa
     3181 gtccttgtct ttaatgattt tcaggagatc gtgatacgtt cccagggatg cgttgaagcg
     3241 atcctccact ccgctgattt caacagagtc gaaacattca atctttttga aatagtcttc
     3301 tttgagctgt ttcacggtaa ctttccggtt cgtcttgaag aggaggtcca cgatagcttt
     3361 cttctgctct ccagacagga atgctggctt tctcatccct tctgtgacgt atttgacctt
     3421 ggtgagctcg ttataaactg tgaagtactc gtacagcaga gagtgtttag gaagcacctt
     3481 ttcgttaggc agatttttat caaagttagt catcctttcg atgaaggact gggcagaggc
     3541 ccccttatcc acgacttcct cgaagttcca gggagtgatg gtctcttctg atttgcgagt
     3601 catccacgcg aatctggaat ttccccgggc gagggggcct acatagtagg gtatccgaaa
     3661 tgtgaggatt ttctcaatct tttccctgtt atctttcaaa aaggggtaga aatcctcttg
     3721 ccgcctgagg atagcgtgca gttcgcccag gtgaatctgg tgggggatgc ttccattgtc
     3781 gaaagtgcgc tgtttgcgca acagatcttc tctgttaagc tttaccagca gctcctcggt
     3841 gccgtccatt ttttccaaga tgggcttaat aaatttgtaa aattcctcct ggcttgctcc
     3901 gccgtcaatg tatccggcgt agccattttt agactgatcg aagaaaattt ccttgtactt
     3961 ctcaggcagt tgctgtctga caagggcctt cagcaaagtc aagtcttggt ggtgctcatc
     4021 atagcgcttg atcatactag cgctcagcgg agctttggtg atctccgtgt tcactcgcag
     4081 aatatcactc agcagaatgg cgtctgacag gttctttgcc gccaaaaaaa ggtctgcgta
     4141 ctggtcgccg atctgggcca gcagattgtc gagatcatca tcgtaggtgt ctttgctcag
     4201 ttgaagcttg gcatcttcgg ccaggtcgaa gttagattta aagttggggg tcagcccgag
     4261 tgacagggcg ataagattac caaacaggcc gttcttcttc tccccaggga gctgtgcgat
     4321 gaggttttcg agccgccggg atttggacag cctagcgctc aggattgctt tggcgtcaac
     4381 tccggatgcg ttgatcgggt tctcttcgaa aagctgattg taagtctgaa ccagttggat
     4441 aaagagtttg tcaacatcgc tgttgtctgg gttcaggtcc ccctcgatga ggaagtgtcc
     4501 ccgaaatttg atcatatgcg ccagcgcgag atagatcaac cgcaagtcag ccttatcagt
     4561 actgtctaca agcttcttcc tcagatgata tatggttggg tacttttcat ggtacgccac
     4621 ctcgtccacg atattgccaa agattgggtg gcgctcgtgc tttttatcct cctccaccaa
     4681 aaaggactcc tccagcctat ggaagaaaga gtcatccacc ttagccatct cattactaaa
     4741 gatctcctgc aggtagcaga tccgattctt tctgcgggta tatctgcgcc gtgctgttct
     4801 tttgagccgc gtggcttcgg ccgtctcccc ggagtcgaac aggagggcgc caatgaggtt
     4861 cttctttatg ctgtggcgat cggtattgcc cagaactttg aattttttgc tcggcacctt
     4921 gtactcgtcc gtaatgacgg cccagccgac gctgtttgtg ccgatatcga gcccaatgga
     4981 gtacttcttg tcctgcggtt agtactgcaa aaagtgctgg tcggatgacg tggcgtcttg
     5041 tgtcgatggg aatgggacca cacccggccg tgtgctcaaa gcccgtggca attgctgctg
     5101 cctctgaaac tcaccatttt gaatgattct tatactcaga aggaaatgct taacgatttc
     5161 gggtgtgagt tgacaaggag agagagaaaa gaagaggaaa ggtaattcgg ggacggtggt
     5221 cttttatacc cttggctaaa gtcccaacca caaagcaaaa aaattttcag tagtctattt
     5281 tgcgtccggc atgggttacc cggatggcca gacaaagaaa ctagtacaaa gtctgaacaa
     5341 gcgtagattc cagactgcag taccctacgc ccttaacggc aagtgtggga accgggggag
     5401 gtttgatatg tggggtgaag ggggctctcg ccggggttgg gcccgctact gggtcaattt
     5461 ggggtcaatt ggggcaattg gggctgtttt ttgggacaca aatacgccgc caacccggtc
     5521 tctgtcgaca aacccaaaag ggccgaaggc tgggccgagg ctctgagatg gatgagatag
     5581 ctggggaaaa caggtgagta aatgattagt aatcagacac ccaggcagat ccgatacacc
     5641 ggtatgattg catcacgtga cttcattgct cctccacaga gagattagtc atgctctttt
     5701 tcagaatcgc aaacccttcc cccacttact taacccctca agctcaactg aaacagtcac
     5761 tcactgtaga tgtgctccga aggcccagga acaatgtggg gtaggaaaat attatgtcta
     5821 tacgagacga atacggtcac cgactctaat gtaagtgata ccatgtatag tagtgtcttc
     5881 tggtacagaa ctccaagtaa tgtgtactta atcccctgta atacaatacc gtgctctcat
     5941 tctcttacgg tatcattaca ttcatctata tacagccaat gggcaatgcg tgtgctgagc
     6001 tcagcacgtt aaattatcga ccggccagtg gtcccattcg ccatgccgaa gcatgttgcc
     6061 cagccggcgc cagcgaggag gctgggacca tgccggccaa aagcaccgac tcggtgccac
     6121 tttttcaagt tgataacgga ctagccttat tttaacttgc tatttctagc tctaaaaccg
     6181 atggcgtcct gatccagtga cgagcttact cgtttcgtcc tcacggactc atcagactgg
     6241 actgcggtta gtactgcaaa aagtgctggt cggatgacgt ggcgtcttgt gtcgatggga
     6301 atgggaccac acccggccgt gtgctcaaag cccgtggcaa ttgctgctgc ctctgaaact
     6361 caccattttg aatgattctt atactcagaa ggaaatgctt aacgatttcg ggtgtgagtt
     6421 gacaaggaga gagagaaaag aagaggaaag gtaattcggg gacggtggtc ttttataccc
     6481 ttggctaaag tcccaaccac aaagcaaaaa aattttcagt agtctatttt gcgtccggca
     6541 tgggttaccc ggatggccag acaaagaaac tagtacaaag tctgaacaag cgtagattcc
     6601 agactgcagt accctacgcc cttaacggca agtgtgggaa ccgggggagg tttgatatgt
     6661 ggggtgaagg gggctctcgc cggggttggg cccgctactg ggtcaatttg gggtcaattg
     6721 gggcaattgg ggctgttttt tgggacacaa atacgccgcc aacccggtct ctgtttaaac
     6781 aaacaagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca
     6841 caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag
     6901 tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt
     6961 cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc
     7021 gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg
     7081 tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa
     7141 agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg
     7201 cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga
     7261 ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg
     7321 tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg
     7381 gaagcgtggc gctttctcaa tgctcacgct gtaggtatct cagttcggtg taggtcgttc
     7441 gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg
     7501 gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca
     7561 ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt
     7621 ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg ctgaagccag
     7681 ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg
     7741 gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc
     7801 ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt
     7861 tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt
     7921 ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca
     7981 gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg
     8041 tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac
     8101 cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg
     8161 ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc
     8221 gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta
     8281 caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac
     8341 gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc
     8401 ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac
     8461 tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact
     8521 caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa
     8581 tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt
     8641 cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca
     8701 ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa
     8761 aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac
     8821 tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg
     8881 gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc
     8941 gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc tataaaaata
     9001 ggcgtatcac gaggcccaga tcctctagag tcgaagcggc cgctatgtct gataaaagga
     9061 tgtaacatag gcaagctgct cgtgagtgtt gagtacgaac cttagatcca aatcacccgc
     9121 acccacggat atacttgctt gaatatacag tagtatgcgg ccgcttcgac accatatcat
     9181 ataaaactaa caatgcattg cttattacga agactacccg ttgctatctc cacaccgtta
     9241 tctccacggt ccaaaggctg ctcaatgtgc tgcatacgta acgtggggtg caaccttgag
     9301 cacatagtac ttttccgaaa accggcgata attaagtgtg cactccaact tttcacactg
     9361 agcgtaaaat gtggagaaga aatcggcact aaaaagtcag gtagactgga aaatgcgcca
     9421 tgaaatgaat atctcttgct acagtaatgc ccagcatcga ggggtattgt gtcaccaaca
     9481 ctatagtggc agctgaagcg ctcgtgattg tagtatgagt ctttattggt gatgggaaga
     9541 gttcactcaa tattctcgtt actgccaaaa caccacggta atcggccaga caccatggat
     9601 gtagatcacc aagcctgtga atgttattcg agctaaaatg cacatggttg gtgaaaggag
     9661 tagttgctgt cgaattccgt cgtcgcctga gtcatcattt atttaccagt tggccacaaa
     9721 cccttgacga tctcgtatgt cccctccgac atactcccgg ccggctgggg tacgttcgat
     9781 agcgctatcg gcatcgacaa ggtttgggtc cctagccgat accgcactac ctgagtcaca
     9841 atcttcggag gtttagtctt ccacatagca cgggcaaaag tgcgtatata tacaagagcg
     9901 tttgccagcc acagattttc actccacaca ccacatcaca catacaacca cacacatcca
     9961 caatggaacc cgaaactaag aagaccaaga ctgactccaa gaagattgtt cttctcggcg
    10021 gcgacttctg tggccccgag gtgattgccg aggccgtcaa ggtgctcaag tctgttgctg
    10081 aggcctccgg caccgagttt gtgtttgagg accgactcat tggaggagct gccattgaga
    10141 aggagggcga gcccatcacc gacgctactc tcgacatctg ccgaaaggct gactctatta
    10201 tgctcggtgc tgtcggaggc gctgccaaca ccgtatggac cactcccgac ggacgaaccg
    10261 acgtgcgacc cgagcagggt ctcctcaagc tgcgaaagga cctgaacctg tacgccaacc
    10321 tgcgaccctg ccagctgctg tcgcccaagc tcgccgatct ctcccccatc cgaaacgttg
    10381 agggcaccga cttcatcatt gtccgagagc tcgtcggagg tatctacttt ggagagcgaa
    10441 aggaggatga cggatctggc gtcgcttccg acaccgagac ctactccgtt cctgaggttg
    10501 agcgaattgc ccgaatggcc gccttcctgg cccttcagca caacccccct cttcccgtgt
    10561 ggtctcttga caaggccaac gtgctggcct cctctcgact ttggcgaaag actgtcactc
    10621 gagtcctcaa ggacgaattc ccccagctcg agctcaacca ccagctgatc gactcggccg
    10681 ccatgatcct catcaagcag ccctccaaga tgaatggtat catcatcacc accaacatgt
    10741 ttggcgatat catctccgac gaggcctccg tcatccccgg ttctctgggt ctgctgccct
    10801 ccgcctctct ggcttctctg cccgacacca acgaggcgtt cggtctgtac gagccctgtc
    10861 acggatctgc ccccgatctc ggcaagcaga aggtcaaccc cattgccacc attctgtctg
    10921 ccgccatgat gctcaagttc tctcttaaca tgaagcccgc cggtgacgct gttgaggctg
    10981 ccgtcaagga gtccgtcgag gctggtatca ctaccgccga tatcggaggc tcttcctcca
    11041 cctccgaggt cggagacttg ttgccaacaa ggtcaaggag ctgctcaaga aggagtaagt
    11101 cgtttctacg acgcattgat ggaaggagca aactgacgcg cctgcgggtt ggtctaccgg
    11161 cagggtccgc tagtgtataa gactctataa aaagggccct gccctgctaa tgaaatgatg
    11221 atttataatt taccggtgta gcaaccttga ctagaagaag cagattgggt gtgtttgtag
    11281 tggaggacag tggtacgttt tggaaacagt cttcttgaaa gtgtcttgtc tacagtatat
    11341 tcactcataa cctcaatagc caagggtgta gtcggtttat taaaggaagg gagttgtggc
    11401 tgatgtggat agatatcttt aagctggcga ctgcacccaa cgagtgtggt ggtagcttgt
    11461 tactgtatat tcggtaagat atattttgtg gggttttagt ggtgtttggt aggttagtgc
    11521 ttggtatatg agttgtaggc atgacaattt ggaaaggggt ggactttggg aatattgtgg
    11581 gatttcaata ccttagtttg tacagggtaa ttgttacaaa tgatacaaag aactgtattt
    11641 cttttcattt gttttaattg gttgtatatc aagtccgtta gacgagctca gtgccatggc
    11701 ttttggcact gtatttcatt tttagaggta cactacattc agtgaggtat ggtaaggttg
    11761 agggcataat gaaggcacct tgtactgaca gtcacagacc tctcaccgag aattttatga
    11821 gatatactcg ggttcatttt aggctccgat tcgattcaaa ttattactgt cgaaatcggt
    11881 tgagcatccg ttgatttccg aacagatctg gccctttcgt c
//
