LOCUS       sequence-329703-       13012 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses spCas9 and a gRNA targeting mouse Yap.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13012)
  AUTHORS   Yu M, Peng Z, Qin M, Liu Y, Wang J, Zhang C, Lin J, Dong T, Wang L, 
            Li S, Yang Y, Xu S, Guo W, Zhang X, Shi M, Peng H, Luo X, Zhang H, 
            Zhang L, Li Y, Yang XP, Sun S
  TITLE     Interferon-gamma induces tumor resistance to anti-PD-1 immunotherapy
            by promoting YAP phase separation.
  JOURNAL   Mol Cell. 2021 Feb 13. pii: S1097-2765(21)00010-1. doi: 
            10.1016/j.molcel.2021.01.010.
  PUBMED    33606996
REFERENCE   2  (bases 1 to 13012)
  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: "Mol Cell. 
            2021 Feb 13. pii: S1097-2765(21)00010-1. doi: 
            10.1016/j.molcel.2021.01.010."
COMMENT     Sequence Label: Lenti-CRISPR-v2-gYap-1
FEATURES             Location/Qualifiers
     source          1..13012
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        382..580
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             598..778
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    825..950
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1443..1676
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1861..1905
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2203..2320
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2371..2619
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2371..2391
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2542..2561
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2640..2715
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        2777..2988
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      3007..3016
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3013..7116
                     /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             3013..7116
                     /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             7117..7164
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7165..7188
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             7198..7254
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             7255..7851
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
                     TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
                     QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
                     PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     7748..7768
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    7867..8455
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7920..7940)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8338..8349)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             8527..8760
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(8786..8803)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    8792..9016
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      9062..9490
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9149..9168)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9359..9380
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(9499..9519)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        9504..9833
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      9684..9819
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     9746..9765
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        9881..9928
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             9947..10321
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    10451..10572
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(10488..10507)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     10542..10561
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(10621..10637)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(10621..10637)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(10634..10656)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    10645..10661
                     /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(10669..10675,10676..10693,10694..10699))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    10714..10735
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(10852..10869)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11023..11611)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11103..11122)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(11782..12573,12574..12642))
                     /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     12405..12424
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(12643..12747)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(12822..12841)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag cgcgttttgc ctgtactggg
      601 tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg
      661 cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt
      721 gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt
      781 ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga ggagctctct cgacgcagga
      841 ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg agtacgccaa
      901 aaattttgac tagcggaggc tagaaggaga gagatgggtg cgagagcgtc agtattaagc
      961 gggggagaat tagatcgcga tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat
     1021 ataaattaaa acatatagta tgggcaagca gggagctaga acgattcgca gttaatcctg
     1081 gcctgttaga aacatcagaa ggctgtagac aaatactggg acagctacaa ccatcccttc
     1141 agacaggatc agaagaactt agatcattat ataatacagt agcaaccctc tattgtgtgc
     1201 atcaaaggat agagataaaa gacaccaagg aagctttaga caagatagag gaagagcaaa
     1261 acaaaagtaa gaccaccgca cagcaagcgg ccgctgatct tcagacctgg aggaggagat
     1321 atgagggaca attggagaag tgaattatat aaatataaag tagtaaaaat tgaaccatta
     1381 ggagtagcac ccaccaaggc aaagagaaga gtggtgcaga gagaaaaaag agcagtggga
     1441 ataggagctt tgttccttgg gttcttggga gcagcaggaa gcactatggg cgcagcgtca
     1501 atgacgctga cggtacaggc cagacaatta ttgtctggta tagtgcagca gcagaacaat
     1561 ttgctgaggg ctattgaggc gcaacagcat ctgttgcaac tcacagtctg gggcatcaag
     1621 cagctccagg caagaatcct ggctgtggaa agatacctaa aggatcaaca gctcctgggg
     1681 atttggggtt gctctggaaa actcatttgc accactgctg tgccttggaa tgctagttgg
     1741 agtaataaat ctctggaaca gatttggaat cacacgacct ggatggagtg ggacagagaa
     1801 attaacaatt acacaagctt aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa
     1861 aagaatgaac aagaattatt ggaattagat aaatgggcaa gtttgtggaa ttggtttaac
     1921 ataacaaatt ggctgtggta tataaaatta ttcataatga tagtaggagg cttggtaggt
     1981 ttaagaatag tttttgctgt actttctata gtgaatagag ttaggcaggg atattcacca
     2041 ttatcgtttc agacccacct cccaaccccg aggggacccg acaggcccga aggaatagaa
     2101 gaagaaggtg gagagagaga cagagacaga tccattcgat tagtgaacgg atcggcactg
     2161 cgtgcgccaa ttctgcagac aaatggcagt attcatccac aattttaaaa gaaaaggggg
     2221 gattgggggg tacagtgcag gggaaagaat agtagacata atagcaacag acatacaaac
     2281 taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt cgggtttatt acagggacag
     2341 cagagatcca gtttggttaa ttaaggtacc gagggcctat ttcccatgat tccttcatat
     2401 ttgcatatac gatacaaggc tgttagagag ataattagaa ttaatttgac tgtaaacaca
     2461 aagatattag tacaaaatac gtgacgtaga aagtaataat ttcttgggta gtttgcagtt
     2521 ttaaaattat gttttaaaat ggactatcat atgcttaccg taacttgaaa gtatttcgat
     2581 ttcttggctt tatatatctt gtggaaagga cgaaacaccc cgccgtccgt gtctccggcg
     2641 ttttagagct agaaatagca agttaaaata aggctagtcc gttatcaact tgaaaaagtg
     2701 gcaccgagtc ggtgcttttt tgaattcgct agctaggtct tgaaaggagt gggaattggc
     2761 tccggtgccc gtcagtgggc agagcgcaca tcgcccacag tccccgagaa gttgggggga
     2821 ggggtcggca attgatccgg tgcctagaga aggtggcgcg gggtaaactg ggaaagtgat
     2881 gtcgtgtact ggctccgcct ttttcccgag ggtgggggag aaccgtatat aagtgcagta
     2941 gtcgccgtga acgttctttt tcgcaacggg tttgccgcca gaacacagga ccggttctag
     3001 agcgctgcca ccatggacaa gaagtacagc atcggcctgg acatcggcac caactctgtg
     3061 ggctgggccg tgatcaccga cgagtacaag gtgcccagca agaaattcaa ggtgctgggc
     3121 aacaccgacc ggcacagcat caagaagaac ctgatcggag ccctgctgtt cgacagcggc
     3181 gaaacagccg aggccacccg gctgaagaga accgccagaa gaagatacac cagacggaag
     3241 aaccggatct gctatctgca agagatcttc agcaacgaga tggccaaggt ggacgacagc
     3301 ttcttccaca gactggaaga gtccttcctg gtggaagagg ataagaagca cgagcggcac
     3361 cccatcttcg gcaacatcgt ggacgaggtg gcctaccacg agaagtaccc caccatctac
     3421 cacctgagaa agaaactggt ggacagcacc gacaaggccg acctgcggct gatctatctg
     3481 gccctggccc acatgatcaa gttccggggc cacttcctga tcgagggcga cctgaacccc
     3541 gacaacagcg acgtggacaa gctgttcatc cagctggtgc agacctacaa ccagctgttc
     3601 gaggaaaacc ccatcaacgc cagcggcgtg gacgccaagg ccatcctgtc tgccagactg
     3661 agcaagagca gacggctgga aaatctgatc gcccagctgc ccggcgagaa gaagaatggc
     3721 ctgttcggaa acctgattgc cctgagcctg ggcctgaccc ccaacttcaa gagcaacttc
     3781 gacctggccg aggatgccaa actgcagctg agcaaggaca cctacgacga cgacctggac
     3841 aacctgctgg cccagatcgg cgaccagtac gccgacctgt ttctggccgc caagaacctg
     3901 tccgacgcca tcctgctgag cgacatcctg agagtgaaca ccgagatcac caaggccccc
     3961 ctgagcgcct ctatgatcaa gagatacgac gagcaccacc aggacctgac cctgctgaaa
     4021 gctctcgtgc ggcagcagct gcctgagaag tacaaagaga ttttcttcga ccagagcaag
     4081 aacggctacg ccggctacat tgacggcgga gccagccagg aagagttcta caagttcatc
     4141 aagcccatcc tggaaaagat ggacggcacc gaggaactgc tcgtgaagct gaacagagag
     4201 gacctgctgc ggaagcagcg gaccttcgac aacggcagca tcccccacca gatccacctg
     4261 ggagagctgc acgccattct gcggcggcag gaagattttt acccattcct gaaggacaac
     4321 cgggaaaaga tcgagaagat cctgaccttc cgcatcccct actacgtggg ccctctggcc
     4381 aggggaaaca gcagattcgc ctggatgacc agaaagagcg aggaaaccat caccccctgg
     4441 aacttcgagg aagtggtgga caagggcgct tccgcccaga gcttcatcga gcggatgacc
     4501 aacttcgata agaacctgcc caacgagaag gtgctgccca agcacagcct gctgtacgag
     4561 tacttcaccg tgtataacga gctgaccaaa gtgaaatacg tgaccgaggg aatgagaaag
     4621 cccgccttcc tgagcggcga gcagaaaaag gccatcgtgg acctgctgtt caagaccaac
     4681 cggaaagtga ccgtgaagca gctgaaagag gactacttca agaaaatcga gtgcttcgac
     4741 tccgtggaaa tctccggcgt ggaagatcgg ttcaacgcct ccctgggcac ataccacgat
     4801 ctgctgaaaa ttatcaagga caaggacttc ctggacaatg aggaaaacga ggacattctg
     4861 gaagatatcg tgctgaccct gacactgttt gaggacagag agatgatcga ggaacggctg
     4921 aaaacctatg cccacctgtt cgacgacaaa gtgatgaagc agctgaagcg gcggagatac
     4981 accggctggg gcaggctgag ccggaagctg atcaacggca tccgggacaa gcagtccggc
     5041 aagacaatcc tggatttcct gaagtccgac ggcttcgcca acagaaactt catgcagctg
     5101 atccacgacg acagcctgac ctttaaagag gacatccaga aagcccaggt gtccggccag
     5161 ggcgatagcc tgcacgagca cattgccaat ctggccggca gccccgccat taagaagggc
     5221 atcctgcaga cagtgaaggt ggtggacgag ctcgtgaaag tgatgggccg gcacaagccc
     5281 gagaacatcg tgatcgaaat ggccagagag aaccagacca cccagaaggg acagaagaac
     5341 agccgcgaga gaatgaagcg gatcgaagag ggcatcaaag agctgggcag ccagatcctg
     5401 aaagaacacc ccgtggaaaa cacccagctg cagaacgaga agctgtacct gtactacctg
     5461 cagaatgggc gggatatgta cgtggaccag gaactggaca tcaaccggct gtccgactac
     5521 gatgtggacc atatcgtgcc tcagagcttt ctgaaggacg actccatcga caacaaggtg
     5581 ctgaccagaa gcgacaagaa ccggggcaag agcgacaacg tgccctccga agaggtcgtg
     5641 aagaagatga agaactactg gcggcagctg ctgaacgcca agctgattac ccagagaaag
     5701 ttcgacaatc tgaccaaggc cgagagaggc ggcctgagcg aactggataa ggccggcttc
     5761 atcaagagac agctggtgga aacccggcag atcacaaagc acgtggcaca gatcctggac
     5821 tcccggatga acactaagta cgacgagaat gacaagctga tccgggaagt gaaagtgatc
     5881 accctgaagt ccaagctggt gtccgatttc cggaaggatt tccagtttta caaagtgcgc
     5941 gagatcaaca actaccacca cgcccacgac gcctacctga acgccgtcgt gggaaccgcc
     6001 ctgatcaaaa agtaccctaa gctggaaagc gagttcgtgt acggcgacta caaggtgtac
     6061 gacgtgcgga agatgatcgc caagagcgag caggaaatcg gcaaggctac cgccaagtac
     6121 ttcttctaca gcaacatcat gaactttttc aagaccgaga ttaccctggc caacggcgag
     6181 atccggaagc ggcctctgat cgagacaaac ggcgaaaccg gggagatcgt gtgggataag
     6241 ggccgggatt ttgccaccgt gcggaaagtg ctgagcatgc cccaagtgaa tatcgtgaaa
     6301 aagaccgagg tgcagacagg cggcttcagc aaagagtcta tcctgcccaa gaggaacagc
     6361 gataagctga tcgccagaaa gaaggactgg gaccctaaga agtacggcgg cttcgacagc
     6421 cccaccgtgg cctattctgt gctggtggtg gccaaagtgg aaaagggcaa gtccaagaaa
     6481 ctgaagagtg tgaaagagct gctggggatc accatcatgg aaagaagcag cttcgagaag
     6541 aatcccatcg actttctgga agccaagggc tacaaagaag tgaaaaagga cctgatcatc
     6601 aagctgccta agtactccct gttcgagctg gaaaacggcc ggaagagaat gctggcctct
     6661 gccggcgaac tgcagaaggg aaacgaactg gccctgccct ccaaatatgt gaacttcctg
     6721 tacctggcca gccactatga gaagctgaag ggctcccccg aggataatga gcagaaacag
     6781 ctgtttgtgg aacagcacaa gcactacctg gacgagatca tcgagcagat cagcgagttc
     6841 tccaagagag tgatcctggc cgacgctaat ctggacaaag tgctgtccgc ctacaacaag
     6901 caccgggata agcccatcag agagcaggcc gagaatatca tccacctgtt taccctgacc
     6961 aatctgggag cccctgccgc cttcaagtac tttgacacca ccatcgaccg gaagaggtac
     7021 accagcacca aagaggtgct ggacgccacc ctgatccacc agagcatcac cggcctgtac
     7081 gagacacgga tcgacctgtc tcagctggga ggcgacaagc gacctgccgc cacaaagaag
     7141 gctggacagg ctaagaagaa gaaagattac aaagacgatg acgataaggg atccggcgca
     7201 acaaacttct ctctgctgaa acaagccgga gatgtcgaag agaatcctgg accgaccgag
     7261 tacaagccca cggtgcgcct cgccacccgc gacgacgtcc ccagggccgt acgcaccctc
     7321 gccgccgcgt tcgccgacta ccccgccacg cgccacaccg tcgatccgga ccgccacatc
     7381 gagcgggtca ccgagctgca agaactcttc ctcacgcgcg tcgggctcga catcggcaag
     7441 gtgtgggtcg cggacgacgg cgccgcggtg gcggtctgga ccacgccgga gagcgtcgaa
     7501 gcgggggcgg tgttcgccga gatcggcccg cgcatggccg agttgagcgg ttcccggctg
     7561 gccgcgcagc aacagatgga aggcctcctg gcgccgcacc ggcccaagga gcccgcgtgg
     7621 ttcctggcca ccgtcggagt ctcgcccgac caccagggca agggtctggg cagcgccgtc
     7681 gtgctccccg gagtggaggc ggccgagcgc gccggggtgc ccgccttcct ggagacctcc
     7741 gcgccccgca acctcccctt ctacgagcgg ctcggcttca ccgtcaccgc cgacgtcgag
     7801 gtgcccgaag gaccgcgcac ctggtgcatg acccgcaagc ccggtgcctg aacgcgttaa
     7861 gtcgacaatc aacctctgga ttacaaaatt tgtgaaagat tgactggtat tcttaactat
     7921 gttgctcctt ttacgctatg tggatacgct gctttaatgc ctttgtatca tgctattgct
     7981 tcccgtatgg ctttcatttt ctcctccttg tataaatcct ggttgctgtc tctttatgag
     8041 gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc
     8101 cccactggtt ggggcattgc caccacctgt cagctccttt ccgggacttt cgctttcccc
     8161 ctccctattg ccacggcgga actcatcgcc gcctgccttg cccgctgctg gacaggggct
     8221 cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga aatcatcgtc ctttccttgg
     8281 ctgctcgcct gtgttgccac ctggattctg cgcgggacgt ccttctgcta cgtcccttcg
     8341 gccctcaatc cagcggacct tccttcccgc ggcctgctgc cggctctgcg gcctcttccg
     8401 cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt gggccgcctc cccgcgtcga
     8461 ctttaagacc aatgacttac aaggcagctg tagatcttag ccacttttta aaagaaaagg
     8521 ggggactgga agggctaatt cactcccaac gaagacaaga tctgcttttt gcttgtactg
     8581 ggtctctctg gttagaccag atctgagcct gggagctctc tggctaacta gggaacccac
     8641 tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt
     8701 gtgactctgg taactagaga tccctcagac ccttttagtc agtgtggaaa atctctagca
     8761 gggcccgttt aaacccgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt
     8821 gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc
     8881 taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt
     8941 ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca tgctggggat
     9001 gcggtgggct ctatggcttc tgaggcggaa agaaccagct ggggctctag ggggtatccc
     9061 cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc
     9121 gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc
     9181 acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt
     9241 agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg
     9301 ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt
     9361 ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta
     9421 taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt
     9481 aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg tggaaagtcc ccaggctccc
     9541 cagcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg tgtggaaagt
     9601 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
     9661 tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc gcccattctc
     9721 cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc tctgcctctg
     9781 agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc aaaaagctcc
     9841 cgggagcttg tatatccatt ttcggatctg atcagcacgt gttgacaatt aatcatcggc
     9901 atagtatatc ggcatagtat aatacgacaa ggtgaggaac taaaccatgg ccaagttgac
     9961 cagtgccgtt ccggtgctca ccgcgcgcga cgtcgccgga gcggtcgagt tctggaccga
    10021 ccggctcggg ttctcccggg acttcgtgga ggacgacttc gccggtgtgg tccgggacga
    10081 cgtgaccctg ttcatcagcg cggtccagga ccaggtggtg ccggacaaca ccctggcctg
    10141 ggtgtgggtg cgcggcctgg acgagctgta cgccgagtgg tcggaggtcg tgtccacgaa
    10201 cttccgggac gcctccgggc cggccatgac cgagatcggc gagcagccgt gggggcggga
    10261 gttcgccctg cgcgacccgg ccggcaactg cgtgcacttc gtggccgagg agcaggactg
    10321 acacgtgcta cgagatttcg attccaccgc cgccttctat gaaaggttgg gcttcggaat
    10381 cgttttccgg gacgccggct ggatgatcct ccagcgcggg gatctcatgc tggagttctt
    10441 cgcccacccc aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac
    10501 aaatttcaca aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat
    10561 caatgtatct tatcatgtct gtataccgtc gacctctagc tagagcttgg cgtaatcatg
    10621 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
    10681 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
    10741 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat
    10801 cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac
    10861 tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt
    10921 aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca
    10981 gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc
    11041 ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact
    11101 ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct
    11161 gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag
    11221 ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca
    11281 cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa
    11341 cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc
    11401 gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag
    11461 aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg
    11521 tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca
    11581 gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc
    11641 tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag
    11701 gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata
    11761 tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat
    11821 ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg
    11881 ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc
    11941 tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc
    12001 aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc
    12061 gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc
    12121 gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc
    12181 ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa
    12241 gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat
    12301 gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata
    12361 gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca
    12421 tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag
    12481 gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc
    12541 agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc
    12601 aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata
    12661 ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta
    12721 gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcga
    12781 cggatcggga gatctcccga tcccctatgg tgcactctca gtacaatctg ctctgatgcc
    12841 gcatagttaa gccagtatct gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg
    12901 agcaaaattt aagctacaac aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt
    12961 agggttaggc gttttgcgct gcttcgcgat gtacgggcca gatatacgcg tt
//
