LOCUS Exported 15261 bp ds-DNA circular SYN 15-JUN-2022 DEFINITION Variant of lentiCRISPRv2 that confers hygromycin resistance. ACCESSION . VERSION . KEYWORDS lentiCRISPRv2 hygro SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 15261) AUTHORS Stringer BW, Day BW, D'Souza RCJ, Jamieson PR, Ensbey KS, Bruce ZC, Lim YC, Goasdoue K, Offenhauser C, Akgul S, Allan S, Robertson T, Lucas P, Tollesson G, Campbell S, Winter C, Do H, Dobrovic A, Inglis PL, Jeffree RL, Johns TG, Boyd AW TITLE A reference collection of patient-derived cell line and xenograft models of proneural, classical and mesenchymal glioblastoma. JOURNAL Sci Rep. 2019 Mar 20;9(1):4902. doi: 10.1038/s41598-019-41277-z. PUBMED 30894629 REFERENCE 2 (bases 1 to 15261) AUTHORS . TITLE Direct Submission JOURNAL Exported Jun 15, 2022 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..15261 /organism="synthetic DNA construct" /mol_type="other DNA" primer_bind complement(47..66) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" enhancer 242..621 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 623..821 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 776..796 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" LTR 839..1019 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" misc_feature 1066..1191 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 1684..1917 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." CDS 2102..2146 /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 2444..2561 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" promoter 2612..2852 /label=U6 promoter /note="RNA polymerase III promoter for human U6 snRNA" primer_bind 2612..2632 /label=hU6-F /note="Human U6 promoter, forward primer" primer_bind 2783..2802 /label=LKO.1 5' /note="Human U6 promoter, forward primer" misc_RNA 4742..4817 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" promoter 4879..5090 /label=EF-1-alpha core promoter /note="core promoter for human elongation factor EF-1-alpha" regulatory 5109..5118 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 5115..9218 /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 9219..9266 /codon_start=1 /product="bipartite nuclear localization signal from nucleoplasmin" /label=nucleoplasmin NLS /translation="KRPAATKKAGQAKKKK" CDS 9267..9290 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 9300..9356 /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" misc_feature 10353..10941 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(10406..10426) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(10824..10835) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" LTR 11013..11246 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" primer_bind complement(11272..11289) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 11278..11502 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 11548..11976 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(11635..11654) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 11845..11866 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(11985..12005) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 11990..12319 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 12170..12305 /label=SV40 ori /note="SV40 origin of replication" primer_bind 12232..12251 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" promoter 12367..12414 /label=EM7 promoter /note="synthetic bacterial promoter " CDS 12433..12807 /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 12937..13058 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(12974..12993) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 13028..13047 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(13107..13123) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(13107..13123) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(13120..13142) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 13131..13147 /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(13155..13185) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 13200..13221 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(13338..13355) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(13509..14097) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(13589..13608) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(14268..15128) /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 14891..14910 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(15129..15233) /gene="bla" /label=AmpR promoter ORIGIN 1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg 61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt 121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc 181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgcgcgt 241 tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt agttcatagc 301 ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc 361 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg 421 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat 481 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc 541 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta 601 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag 661 cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt 721 tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa 781 atgggcggta ggcgtgtacg gtgggaggtc tatataagca gcgcgttttg cctgtactgg 841 gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact 901 gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg 961 tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag 1021 tggcgcccga acagggactt gaaagcgaaa gggaaaccag aggagctctc tcgacgcagg 1081 actcggcttg ctgaagcgcg cacggcaaga ggcgaggggc ggcgactggt gagtacgcca 1141 aaaattttga ctagcggagg ctagaaggag agagatgggt gcgagagcgt cagtattaag 1201 cgggggagaa ttagatcgcg atgggaaaaa attcggttaa ggccaggggg aaagaaaaaa 1261 tataaattaa aacatatagt atgggcaagc agggagctag aacgattcgc agttaatcct 1321 ggcctgttag aaacatcaga aggctgtaga caaatactgg gacagctaca accatccctt 1381 cagacaggat cagaagaact tagatcatta tataatacag tagcaaccct ctattgtgtg 1441 catcaaagga tagagataaa agacaccaag gaagctttag acaagataga ggaagagcaa 1501 aacaaaagta agaccaccgc acagcaagcg gccgctgatc ttcagacctg gaggaggaga 1561 tatgagggac aattggagaa gtgaattata taaatataaa gtagtaaaaa ttgaaccatt 1621 aggagtagca cccaccaagg caaagagaag agtggtgcag agagaaaaaa gagcagtggg 1681 aataggagct ttgttccttg ggttcttggg agcagcagga agcactatgg gcgcagcgtc 1741 aatgacgctg acggtacagg ccagacaatt attgtctggt atagtgcagc agcagaacaa 1801 tttgctgagg gctattgagg cgcaacagca tctgttgcaa ctcacagtct ggggcatcaa 1861 gcagctccag gcaagaatcc tggctgtgga aagataccta aaggatcaac agctcctggg 1921 gatttggggt tgctctggaa aactcatttg caccactgct gtgccttgga atgctagttg 1981 gagtaataaa tctctggaac agatttggaa tcacacgacc tggatggagt gggacagaga 2041 aattaacaat tacacaagct taatacactc cttaattgaa gaatcgcaaa accagcaaga 2101 aaagaatgaa caagaattat tggaattaga taaatgggca agtttgtgga attggtttaa 2161 cataacaaat tggctgtggt atataaaatt attcataatg atagtaggag gcttggtagg 2221 tttaagaata gtttttgctg tactttctat agtgaataga gttaggcagg gatattcacc 2281 attatcgttt cagacccacc tcccaacccc gaggggaccc gacaggcccg aaggaataga 2341 agaagaaggt ggagagagag acagagacag atccattcga ttagtgaacg gatcggcact 2401 gcgtgcgcca attctgcaga caaatggcag tattcatcca caattttaaa agaaaagggg 2461 ggattggggg gtacagtgca ggggaaagaa tagtagacat aatagcaaca gacatacaaa 2521 ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt tcgggtttat tacagggaca 2581 gcagagatcc agtttggtta attaaggtac cgagggccta tttcccatga ttccttcata 2641 tttgcatata cgatacaagg ctgttagaga gataattaga attaatttga ctgtaaacac 2701 aaagatatta gtacaaaata cgtgacgtag aaagtaataa tttcttgggt agtttgcagt 2761 tttaaaatta tgttttaaaa tggactatca tatgcttacc gtaacttgaa agtatttcga 2821 tttcttggct ttatatatct tgtggaaagg acgaaacacc ggagacggtt gtaaatgagc 2881 acacaaaata cacatgctaa aatattatat tctatgacct ttataaaatc aaccaaaatc 2941 ttctttttaa taactttagt atcaataatt agaattttta tgttcctttt tgcaaacttt 3001 taataaaaat gagcaaaata aaaaaacgct agttttagta actcgcgttg ttttcttcac 3061 ctttaataat agctactcca ccacttgttc ctaagcggtc agctcctgct tcaatcattt 3121 tttgagcatc ttcaaatgtt ctaactccac cagctgcttt aactaaagca ttgtctttaa 3181 caactgactt cattagttta acatcttcaa atgttgcacc tgattttgaa aatcctgttg 3241 atgttttaac aaattctaat ccagcttcaa cagctatttc acaagctttc atgatttctt 3301 cttttgttaa taaacaattt tccataatac atttaacaac atgtgatcca gctgcttttt 3361 ttacagcttt catgtcttct aaaactaatt cataattttt gtcttttaat gcaccaatat 3421 ttaataccat atcaatttct gttgcaccat ctttaattgc ttcagaaact tcgaatgctt 3481 ttgtagctgt tgtgcatgca cctagaggaa aacctacaac atttgttatt cctacatttg 3541 tgccttttaa taattcttta caatagcttg ttcaatatga attaacacaa actgttgcaa 3601 aatcaaattc aattgcttca tcacataatt gtttaatttc agctttcgta gcatcttgtt 3661 ttaataatgt gtgatctata tatttgttta gtttcatttt ttctcctata tattcatttt 3721 taattttaat tctttaataa tttcgtctac tttaacttta gcgttttgaa cagattcacc 3781 aacacctata aaataaattt ttagtttagg ttcagttcca cttgggcgaa cagcaaatca 3841 tgacttatct tctaaataaa attttagtaa gtcttgtcct ggcatattat acattccatc 3901 gatgtagtct tcaacattaa caactttaag tccagcaatt tgagttaagg gtgttgctct 3961 caatgatttc attaatggtt caatttttaa tttcttttct tctggtttaa aattcaagtt 4021 taaagtgaaa gtgtaatatg cacccatttc tttaaataaa tcttctaaat agtctactaa 4081 tgttttattt tgttttttat aaaatcaagc agcctctgct attaatatag aagcttgtat 4141 tccatcttta tctctagctg agtcatcaat tacatatcca taactttctt cataagcaaa 4201 aacaaaattt aatccgttat cttcttcttt agcaatttct ctacccattc atttaaatcc 4261 agttaaagtt tttacaatat taactccata tttttcatga gcgattctat cacccaaatc 4321 acttgttaca aaacttgaat atagagccgg attttttgga atgctattta agcgttttag 4381 atttgataat tttcaatcaa ttaaaattgg tcctgtttga tttccatcta atcttacaaa 4441 atgaccatca tgttttattg ccattccaaa tctgtcagca tctgggtcat tcataataat 4501 aatatctgca tcatgtttaa taccatattc aagcggtatt tttcatgcag gatcaaattc 4561 tggatttgga tttacaacat ttttaaatgt ttcatcttca aatgcatgct cttcaacctc 4621 aataacgtta tatcctgatt cacgtaatat ttttggggta aatttagttc ctgttccatt 4681 aactgcgcta aaaataattt ttaaatcttt tttagcttct tgctcttttt tgtacgtctc 4741 tgttttagag ctagaaatag caagttaaaa taaggctagt ccgttatcaa cttgaaaaag 4801 tggcaccgag tcggtgcttt tttgaattcg ctagctaggt cttgaaagga gtgggaattg 4861 gctccggtgc ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg 4921 gaggggtcgg caattgatcc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg 4981 atgtcgtgta ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag 5041 tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gaccggttct 5101 agagcgctgc caccatggac aagaagtaca gcatcggcct ggacatcggc accaactctg 5161 tgggctgggc cgtgatcacc gacgagtaca aggtgcccag caagaaattc aaggtgctgg 5221 gcaacaccga ccggcacagc atcaagaaga acctgatcgg agccctgctg ttcgacagcg 5281 gcgaaacagc cgaggccacc cggctgaaga gaaccgccag aagaagatac accagacgga 5341 agaaccggat ctgctatctg caagagatct tcagcaacga gatggccaag gtggacgaca 5401 gcttcttcca cagactggaa gagtccttcc tggtggaaga ggataagaag cacgagcggc 5461 accccatctt cggcaacatc gtggacgagg tggcctacca cgagaagtac cccaccatct 5521 accacctgag aaagaaactg gtggacagca ccgacaaggc cgacctgcgg ctgatctatc 5581 tggccctggc ccacatgatc aagttccggg gccacttcct gatcgagggc gacctgaacc 5641 ccgacaacag cgacgtggac aagctgttca tccagctggt gcagacctac aaccagctgt 5701 tcgaggaaaa ccccatcaac gccagcggcg tggacgccaa ggccatcctg tctgccagac 5761 tgagcaagag cagacggctg gaaaatctga tcgcccagct gcccggcgag aagaagaatg 5821 gcctgttcgg aaacctgatt gccctgagcc tgggcctgac ccccaacttc aagagcaact 5881 tcgacctggc cgaggatgcc aaactgcagc tgagcaagga cacctacgac gacgacctgg 5941 acaacctgct ggcccagatc ggcgaccagt acgccgacct gtttctggcc gccaagaacc 6001 tgtccgacgc catcctgctg agcgacatcc tgagagtgaa caccgagatc accaaggccc 6061 ccctgagcgc ctctatgatc aagagatacg acgagcacca ccaggacctg accctgctga 6121 aagctctcgt gcggcagcag ctgcctgaga agtacaaaga gattttcttc gaccagagca 6181 agaacggcta cgccggctac attgacggcg gagccagcca ggaagagttc tacaagttca 6241 tcaagcccat cctggaaaag atggacggca ccgaggaact gctcgtgaag ctgaacagag 6301 aggacctgct gcggaagcag cggaccttcg acaacggcag catcccccac cagatccacc 6361 tgggagagct gcacgccatt ctgcggcggc aggaagattt ttacccattc ctgaaggaca 6421 accgggaaaa gatcgagaag atcctgacct tccgcatccc ctactacgtg ggccctctgg 6481 ccaggggaaa cagcagattc gcctggatga ccagaaagag cgaggaaacc atcaccccct 6541 ggaacttcga ggaagtggtg gacaagggcg cttccgccca gagcttcatc gagcggatga 6601 ccaacttcga taagaacctg cccaacgaga aggtgctgcc caagcacagc ctgctgtacg 6661 agtacttcac cgtgtataac gagctgacca aagtgaaata cgtgaccgag ggaatgagaa 6721 agcccgcctt cctgagcggc gagcagaaaa aggccatcgt ggacctgctg ttcaagacca 6781 accggaaagt gaccgtgaag cagctgaaag aggactactt caagaaaatc gagtgcttcg 6841 actccgtgga aatctccggc gtggaagatc ggttcaacgc ctccctgggc acataccacg 6901 atctgctgaa aattatcaag gacaaggact tcctggacaa tgaggaaaac gaggacattc 6961 tggaagatat cgtgctgacc ctgacactgt ttgaggacag agagatgatc gaggaacggc 7021 tgaaaaccta tgcccacctg ttcgacgaca aagtgatgaa gcagctgaag cggcggagat 7081 acaccggctg gggcaggctg agccggaagc tgatcaacgg catccgggac aagcagtccg 7141 gcaagacaat cctggatttc ctgaagtccg acggcttcgc caacagaaac ttcatgcagc 7201 tgatccacga cgacagcctg acctttaaag aggacatcca gaaagcccag gtgtccggcc 7261 agggcgatag cctgcacgag cacattgcca atctggccgg cagccccgcc attaagaagg 7321 gcatcctgca gacagtgaag gtggtggacg agctcgtgaa agtgatgggc cggcacaagc 7381 ccgagaacat cgtgatcgaa atggccagag agaaccagac cacccagaag ggacagaaga 7441 acagccgcga gagaatgaag cggatcgaag agggcatcaa agagctgggc agccagatcc 7501 tgaaagaaca ccccgtggaa aacacccagc tgcagaacga gaagctgtac ctgtactacc 7561 tgcagaatgg gcgggatatg tacgtggacc aggaactgga catcaaccgg ctgtccgact 7621 acgatgtgga ccatatcgtg cctcagagct ttctgaagga cgactccatc gacaacaagg 7681 tgctgaccag aagcgacaag aaccggggca agagcgacaa cgtgccctcc gaagaggtcg 7741 tgaagaagat gaagaactac tggcggcagc tgctgaacgc caagctgatt acccagagaa 7801 agttcgacaa tctgaccaag gccgagagag gcggcctgag cgaactggat aaggccggct 7861 tcatcaagag acagctggtg gaaacccggc agatcacaaa gcacgtggca cagatcctgg 7921 actcccggat gaacactaag tacgacgaga atgacaagct gatccgggaa gtgaaagtga 7981 tcaccctgaa gtccaagctg gtgtccgatt tccggaagga tttccagttt tacaaagtgc 8041 gcgagatcaa caactaccac cacgcccacg acgcctacct gaacgccgtc gtgggaaccg 8101 ccctgatcaa aaagtaccct aagctggaaa gcgagttcgt gtacggcgac tacaaggtgt 8161 acgacgtgcg gaagatgatc gccaagagcg agcaggaaat cggcaaggct accgccaagt 8221 acttcttcta cagcaacatc atgaactttt tcaagaccga gattaccctg gccaacggcg 8281 agatccggaa gcggcctctg atcgagacaa acggcgaaac cggggagatc gtgtgggata 8341 agggccggga ttttgccacc gtgcggaaag tgctgagcat gccccaagtg aatatcgtga 8401 aaaagaccga ggtgcagaca ggcggcttca gcaaagagtc tatcctgccc aagaggaaca 8461 gcgataagct gatcgccaga aagaaggact gggaccctaa gaagtacggc ggcttcgaca 8521 gccccaccgt ggcctattct gtgctggtgg tggccaaagt ggaaaagggc aagtccaaga 8581 aactgaagag tgtgaaagag ctgctgggga tcaccatcat ggaaagaagc agcttcgaga 8641 agaatcccat cgactttctg gaagccaagg gctacaaaga agtgaaaaag gacctgatca 8701 tcaagctgcc taagtactcc ctgttcgagc tggaaaacgg ccggaagaga atgctggcct 8761 ctgccggcga actgcagaag ggaaacgaac tggccctgcc ctccaaatat gtgaacttcc 8821 tgtacctggc cagccactat gagaagctga agggctcccc cgaggataat gagcagaaac 8881 agctgtttgt ggaacagcac aagcactacc tggacgagat catcgagcag atcagcgagt 8941 tctccaagag agtgatcctg gccgacgcta atctggacaa agtgctgtcc gcctacaaca 9001 agcaccggga taagcccatc agagagcagg ccgagaatat catccacctg tttaccctga 9061 ccaatctggg agcccctgcc gccttcaagt actttgacac caccatcgac cggaagaggt 9121 acaccagcac caaagaggtg ctggacgcca ccctgatcca ccagagcatc accggcctgt 9181 acgagacacg gatcgacctg tctcagctgg gaggcgacaa gcgacctgcc gccacaaaga 9241 aggctggaca ggctaagaag aagaaagatt acaaagacga tgacgataag ggatccggcg 9301 caacaaactt ctctctgctg aaacaagccg gagatgtcga agagaatcct ggaccgaaaa 9361 agcctgaact caccgcgacg tctgtcgaga agtttctgat cgaaaagttc gacagcgtgt 9421 ccgacctgat gcagctctcg gagggcgaag aatctcgtgc tttcagcttc gatgtaggag 9481 ggcgtggata tgtcctgcgg gtaaatagct gcgccgatgg tttctacaaa gatcgttatg 9541 tttatcggca ctttgcatcg gccgcgctcc cgattccgga agtgcttgac attggggaat 9601 tcagcgagag cctgacctat tgcatctccc gccgtgcaca gggtgtcacg ttgcaagacc 9661 tgcctgaaac cgaactgccc gctgttctgc agccggtcgc ggaggccatg gatgcgatcg 9721 ctgcggccga tcttagccag acgagcgggt tcggcccatt cggaccgcaa ggaatcggtc 9781 aatacactac atggcgtgat ttcatatgcg cgattgctga tccccatgtg tatcactggc 9841 aaactgtgat ggacgacacc gtcagtgcgt ccgtcgcgca ggctctcgat gagctgatgc 9901 tttgggccga ggactgcccc gaagtccggc acctcgtgca cgcggatttc ggctccaaca 9961 atgtcctgac ggacaatggc cgcataacag cggtcattga ctggagcgag gcgatgttcg 10021 gggattccca atacgaggtc gccaacatct tcttctggag gccgtggttg gcttgtatgg 10081 agcagcagac gcgctacttc gagcggaggc atccggagct tgcaggatcg ccgcggctcc 10141 gggcgtatat gctccgcatt ggtcttgacc aactctatca gagcttggtt gacggcaatt 10201 tcgatgatgc agcttgggcg cagggtcgat gcgacgcaat cgtccgatcc ggagccggga 10261 ctgtcgggcg tacacaaatc gcccgcagaa gcgcggccgt ctggaccgat ggctgtgtag 10321 aagtacttcg ccgatagacg cgttaagtcg acaatcaacc tctggattac aaaatttgtg 10381 aaagattgac tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt 10441 taatgccttt gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata 10501 aatcctggtt gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg 10561 tgtgcactgt gtttgctgac gcaaccccca ctggttgggg cattgccacc acctgtcagc 10621 tcctttccgg gactttcgct ttccccctcc ctattgccac ggcggaactc atcgccgcct 10681 gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttgt 10741 cggggaaatc atcgtccttt ccttggctgc tcgcctgtgt tgccacctgg attctgcgcg 10801 ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc ggaccttcct tcccgcggcc 10861 tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg agtcggatct 10921 ccctttgggc cgcctccccg cgtcgacttt aagaccaatg acttacaagg cagctgtaga 10981 tcttagccac tttttaaaag aaaagggggg actggaaggg ctaattcact cccaacgaag 11041 acaagatctg ctttttgctt gtactgggtc tctctggtta gaccagatct gagcctggga 11101 gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct 11161 tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt 11221 ttagtcagtg tggaaaatct ctagcagggc ccgtttaaac ccgctgatca gcctcgactg 11281 tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg 11341 aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga 11401 gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg 11461 aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa 11521 ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg 11581 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt 11641 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc 11701 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg 11761 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga 11821 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc 11881 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa 11941 aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt 12001 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa 12061 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag 12121 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct 12181 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc 12241 agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg 12301 aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca 12361 gcacgtgttg acaattaatc atcggcatag tatatcggca tagtataata cgacaaggtg 12421 aggaactaaa ccatggccaa gttgaccagt gccgttccgg tgctcaccgc gcgcgacgtc 12481 gccggagcgg tcgagttctg gaccgaccgg ctcgggttct cccgggactt cgtggaggac 12541 gacttcgccg gtgtggtccg ggacgacgtg accctgttca tcagcgcggt ccaggaccag 12601 gtggtgccgg acaacaccct ggcctgggtg tgggtgcgcg gcctggacga gctgtacgcc 12661 gagtggtcgg aggtcgtgtc cacgaacttc cgggacgcct ccgggccggc catgaccgag 12721 atcggcgagc agccgtgggg gcgggagttc gccctgcgcg acccggccgg caactgcgtg 12781 cacttcgtgg ccgaggagca ggactgacac gtgctacgag atttcgattc caccgccgcc 12841 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag 12901 cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat 12961 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat 13021 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc 13081 tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg 13141 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa 13201 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac 13261 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt 13321 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga 13381 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca 13441 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg 13501 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt 13561 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc 13621 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct 13681 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc 13741 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta 13801 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca 13861 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag 13921 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag 13981 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt 14041 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa 14101 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg 14161 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga 14221 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta 14281 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc 14341 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg 14401 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga 14461 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt 14521 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt 14581 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc 14641 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc 14701 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca 14761 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag 14821 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg 14881 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa 14941 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa 15001 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga 15061 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga 15121 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg 15181 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt 15241 ccccgaaaag tgccacctga c //