LOCUS Exported 13012 bp ds-DNA circular SYN 10-SEP-2021 DEFINITION Lentiviral vector expressing Cas9 and a sgRNA against the human AMBRA1 gene. ACCESSION . VERSION . KEYWORDS lentiCRISPRv2_AMBRA1#1 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 13012) AUTHORS Chaikovsky AC, Li C, Jeng EE, Loebell S, Lee MC, Murray CW, Cheng R, Demeter J, Swaney DL, Chen SH, Newton BW, Johnson JR, Drainas AP, Shue YT, Seoane JA, Srinivasan P, He A, Yoshida A, Hipkins SQ, McCrea E, Poltorack CD, Krogan NJ, Diehl JA, Kong C, Jackson PK, Curtis C, Petrov DA, Bassik MC, Winslow MM, Sage J TITLE The AMBRA1 E3 ligase adaptor regulates the stability of cyclin D. JOURNAL Nature. 2021 Apr;592(7856):794-798. doi: 10.1038/s41586-021-03474-7. Epub 2021 Apr 14. PUBMED 33854239 REFERENCE 2 (bases 1 to 13012) AUTHORS . TITLE Direct Submission JOURNAL Exported Sep 10, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..13012 /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 238..617 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 619..817 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 772..792 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" LTR 835..1015 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" misc_feature 1062..1187 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 1680..1913 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." CDS 2098..2142 /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 2440..2557 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" promoter 2608..2848 /label=U6 promoter /note="RNA polymerase III promoter for human U6 snRNA" primer_bind 2608..2628 /label=hU6-F /note="Human U6 promoter, forward primer" primer_bind 2779..2798 /label=LKO.1 5' /note="Human U6 promoter, forward primer" misc_RNA 2877..2952 /label=gRNA scaffold /note="guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system" promoter 3014..3225 /label=EF-1-alpha core promoter /note="core promoter for human elongation factor EF-1-alpha" regulatory 3244..3253 /regulatory_class="other" /note="vertebrate consensus sequence for strong initiation of translation (Kozak, 1987)" CDS 3250..7353 /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 7354..7401 /codon_start=1 /product="bipartite nuclear localization signal from nucleoplasmin" /label=nucleoplasmin NLS /translation="KRPAATKKAGQAKKKK" CDS 7402..7425 /codon_start=1 /product="FLAG(R) epitope tag, followed by an enterokinase cleavage site" /label=FLAG /translation="DYKDDDDK" CDS 7435..7491 /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 7492..8088 /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 7985..8005 /label=Puro-F /note="Puromycin resistance gene, forward primer" misc_feature 8104..8692 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(8157..8177) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(8575..8586) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" LTR 8764..8997 /label=3' LTR (Delta-U3) /note="self-inactivating 3' long terminal repeat (LTR) from HIV-1" primer_bind complement(9023..9040) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 9029..9253 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 9299..9727 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(9386..9405) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 9596..9617 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(9736..9756) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 9741..10070 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 9921..10056 /label=SV40 ori /note="SV40 origin of replication" primer_bind 9983..10002 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" promoter 10118..10165 /label=EM7 promoter /note="synthetic bacterial promoter " CDS 10184..10558 /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 10688..10809 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(10725..10744) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 10779..10798 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(10858..10874) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(10858..10874) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(10871..10893) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 10882..10898 /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(10906..10936) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 10951..10972 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(11089..11106) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(11260..11848) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(11340..11359) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(12019..12879) /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 12642..12661 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(12880..12984) /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 acgcgttgac 241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat 301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg 361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt 421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag 481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc 541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag 601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt 661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc 721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg 781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct 841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt 901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac 961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc 1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc 1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa 1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg 1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata 1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc 1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga 1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc 1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca 1501 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg 1561 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga 1621 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata 1681 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg 1741 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg 1801 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag 1861 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt 1921 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt 1981 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt 2041 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag 2101 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata 2161 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta 2221 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta 2281 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa 2341 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt 2401 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat 2461 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa 2521 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag 2581 agatccagtt tggttaatta aggtaccgag ggcctatttc ccatgattcc ttcatatttg 2641 catatacgat acaaggctgt tagagagata attagaatta atttgactgt aaacacaaag 2701 atattagtac aaaatacgtg acgtagaaag taataatttc ttgggtagtt tgcagtttta 2761 aaattatgtt ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta tttcgatttc 2821 ttggctttat atatcttgtg gaaaggacga aacaccgata ttatggttca catggggttt 2881 tagagctaga aatagcaagt taaaataagg ctagtccgtt atcaacttga aaaagtggca 2941 ccgagtcggt gcttttttga attcgctagc taggtcttga aaggagtggg aattggctcc 3001 ggtgcccgtc agtgggcaga gcgcacatcg cccacagtcc ccgagaagtt ggggggaggg 3061 gtcggcaatt gatccggtgc ctagagaagg tggcgcgggg taaactggga aagtgatgtc 3121 gtgtactggc tccgcctttt tcccgagggt gggggagaac cgtatataag tgcagtagtc 3181 gccgtgaacg ttctttttcg caacgggttt gccgccagaa cacaggaccg gttctagagc 3241 gctgccacca tggacaagaa gtacagcatc ggcctggaca tcggcaccaa ctctgtgggc 3301 tgggccgtga tcaccgacga gtacaaggtg cccagcaaga aattcaaggt gctgggcaac 3361 accgaccggc acagcatcaa gaagaacctg atcggagccc tgctgttcga cagcggcgaa 3421 acagccgagg ccacccggct gaagagaacc gccagaagaa gatacaccag acggaagaac 3481 cggatctgct atctgcaaga gatcttcagc aacgagatgg ccaaggtgga cgacagcttc 3541 ttccacagac tggaagagtc cttcctggtg gaagaggata agaagcacga gcggcacccc 3601 atcttcggca acatcgtgga cgaggtggcc taccacgaga agtaccccac catctaccac 3661 ctgagaaaga aactggtgga cagcaccgac aaggccgacc tgcggctgat ctatctggcc 3721 ctggcccaca tgatcaagtt ccggggccac ttcctgatcg agggcgacct gaaccccgac 3781 aacagcgacg tggacaagct gttcatccag ctggtgcaga cctacaacca gctgttcgag 3841 gaaaacccca tcaacgccag cggcgtggac gccaaggcca tcctgtctgc cagactgagc 3901 aagagcagac ggctggaaaa tctgatcgcc cagctgcccg gcgagaagaa gaatggcctg 3961 ttcggaaacc tgattgccct gagcctgggc ctgaccccca acttcaagag caacttcgac 4021 ctggccgagg atgccaaact gcagctgagc aaggacacct acgacgacga cctggacaac 4081 ctgctggccc agatcggcga ccagtacgcc gacctgtttc tggccgccaa gaacctgtcc 4141 gacgccatcc tgctgagcga catcctgaga gtgaacaccg agatcaccaa ggcccccctg 4201 agcgcctcta tgatcaagag atacgacgag caccaccagg acctgaccct gctgaaagct 4261 ctcgtgcggc agcagctgcc tgagaagtac aaagagattt tcttcgacca gagcaagaac 4321 ggctacgccg gctacattga cggcggagcc agccaggaag agttctacaa gttcatcaag 4381 cccatcctgg aaaagatgga cggcaccgag gaactgctcg tgaagctgaa cagagaggac 4441 ctgctgcgga agcagcggac cttcgacaac ggcagcatcc cccaccagat ccacctggga 4501 gagctgcacg ccattctgcg gcggcaggaa gatttttacc cattcctgaa ggacaaccgg 4561 gaaaagatcg agaagatcct gaccttccgc atcccctact acgtgggccc tctggccagg 4621 ggaaacagca gattcgcctg gatgaccaga aagagcgagg aaaccatcac cccctggaac 4681 ttcgaggaag tggtggacaa gggcgcttcc gcccagagct tcatcgagcg gatgaccaac 4741 ttcgataaga acctgcccaa cgagaaggtg ctgcccaagc acagcctgct gtacgagtac 4801 ttcaccgtgt ataacgagct gaccaaagtg aaatacgtga ccgagggaat gagaaagccc 4861 gccttcctga gcggcgagca gaaaaaggcc atcgtggacc tgctgttcaa gaccaaccgg 4921 aaagtgaccg tgaagcagct gaaagaggac tacttcaaga aaatcgagtg cttcgactcc 4981 gtggaaatct ccggcgtgga agatcggttc aacgcctccc tgggcacata ccacgatctg 5041 ctgaaaatta tcaaggacaa ggacttcctg gacaatgagg aaaacgagga cattctggaa 5101 gatatcgtgc tgaccctgac actgtttgag gacagagaga tgatcgagga acggctgaaa 5161 acctatgccc acctgttcga cgacaaagtg atgaagcagc tgaagcggcg gagatacacc 5221 ggctggggca ggctgagccg gaagctgatc aacggcatcc gggacaagca gtccggcaag 5281 acaatcctgg atttcctgaa gtccgacggc ttcgccaaca gaaacttcat gcagctgatc 5341 cacgacgaca gcctgacctt taaagaggac atccagaaag cccaggtgtc cggccagggc 5401 gatagcctgc acgagcacat tgccaatctg gccggcagcc ccgccattaa gaagggcatc 5461 ctgcagacag tgaaggtggt ggacgagctc gtgaaagtga tgggccggca caagcccgag 5521 aacatcgtga tcgaaatggc cagagagaac cagaccaccc agaagggaca gaagaacagc 5581 cgcgagagaa tgaagcggat cgaagagggc atcaaagagc tgggcagcca gatcctgaaa 5641 gaacaccccg tggaaaacac ccagctgcag aacgagaagc tgtacctgta ctacctgcag 5701 aatgggcggg atatgtacgt ggaccaggaa ctggacatca accggctgtc cgactacgat 5761 gtggaccata tcgtgcctca gagctttctg aaggacgact ccatcgacaa caaggtgctg 5821 accagaagcg acaagaaccg gggcaagagc gacaacgtgc cctccgaaga ggtcgtgaag 5881 aagatgaaga actactggcg gcagctgctg aacgccaagc tgattaccca gagaaagttc 5941 gacaatctga ccaaggccga gagaggcggc ctgagcgaac tggataaggc cggcttcatc 6001 aagagacagc tggtggaaac ccggcagatc acaaagcacg tggcacagat cctggactcc 6061 cggatgaaca ctaagtacga cgagaatgac aagctgatcc gggaagtgaa agtgatcacc 6121 ctgaagtcca agctggtgtc cgatttccgg aaggatttcc agttttacaa agtgcgcgag 6181 atcaacaact accaccacgc ccacgacgcc tacctgaacg ccgtcgtggg aaccgccctg 6241 atcaaaaagt accctaagct ggaaagcgag ttcgtgtacg gcgactacaa ggtgtacgac 6301 gtgcggaaga tgatcgccaa gagcgagcag gaaatcggca aggctaccgc caagtacttc 6361 ttctacagca acatcatgaa ctttttcaag accgagatta ccctggccaa cggcgagatc 6421 cggaagcggc ctctgatcga gacaaacggc gaaaccgggg agatcgtgtg ggataagggc 6481 cgggattttg ccaccgtgcg gaaagtgctg agcatgcccc aagtgaatat cgtgaaaaag 6541 accgaggtgc agacaggcgg cttcagcaaa gagtctatcc tgcccaagag gaacagcgat 6601 aagctgatcg ccagaaagaa ggactgggac cctaagaagt acggcggctt cgacagcccc 6661 accgtggcct attctgtgct ggtggtggcc aaagtggaaa agggcaagtc caagaaactg 6721 aagagtgtga aagagctgct ggggatcacc atcatggaaa gaagcagctt cgagaagaat 6781 cccatcgact ttctggaagc caagggctac aaagaagtga aaaaggacct gatcatcaag 6841 ctgcctaagt actccctgtt cgagctggaa aacggccgga agagaatgct ggcctctgcc 6901 ggcgaactgc agaagggaaa cgaactggcc ctgccctcca aatatgtgaa cttcctgtac 6961 ctggccagcc actatgagaa gctgaagggc tcccccgagg ataatgagca gaaacagctg 7021 tttgtggaac agcacaagca ctacctggac gagatcatcg agcagatcag cgagttctcc 7081 aagagagtga tcctggccga cgctaatctg gacaaagtgc tgtccgccta caacaagcac 7141 cgggataagc ccatcagaga gcaggccgag aatatcatcc acctgtttac cctgaccaat 7201 ctgggagccc ctgccgcctt caagtacttt gacaccacca tcgaccggaa gaggtacacc 7261 agcaccaaag aggtgctgga cgccaccctg atccaccaga gcatcaccgg cctgtacgag 7321 acacggatcg acctgtctca gctgggaggc gacaagcgac ctgccgccac aaagaaggct 7381 ggacaggcta agaagaagaa agattacaaa gacgatgacg ataagggatc cggcgcaaca 7441 aacttctctc tgctgaaaca agccggagat gtcgaagaga atcctggacc gaccgagtac 7501 aagcccacgg tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg caccctcgcc 7561 gccgcgttcg ccgactaccc cgccacgcgc cacaccgtcg atccggaccg ccacatcgag 7621 cgggtcaccg agctgcaaga actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg 7681 tgggtcgcgg acgacggcgc cgcggtggcg gtctggacca cgccggagag cgtcgaagcg 7741 ggggcggtgt tcgccgagat cggcccgcgc atggccgagt tgagcggttc ccggctggcc 7801 gcgcagcaac agatggaagg cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc 7861 ctggccaccg tcggagtctc gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg 7921 ctccccggag tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga gacctccgcg 7981 ccccgcaacc tccccttcta cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg 8041 cccgaaggac cgcgcacctg gtgcatgacc cgcaagcccg gtgcctgaac gcgttaagtc 8101 gacaatcaac ctctggatta caaaatttgt gaaagattga ctggtattct taactatgtt 8161 gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc tattgcttcc 8221 cgtatggctt tcattttctc ctccttgtat aaatcctggt tgctgtctct ttatgaggag 8281 ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc 8341 actggttggg gcattgccac cacctgtcag ctcctttccg ggactttcgc tttccccctc 8401 cctattgcca cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg 8461 ctgttgggca ctgacaattc cgtggtgttg tcggggaaat catcgtcctt tccttggctg 8521 ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct tctgctacgt cccttcggcc 8581 ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt 8641 cttcgccttc gccctcagac gagtcggatc tccctttggg ccgcctcccc gcgtcgactt 8701 taagaccaat gacttacaag gcagctgtag atcttagcca ctttttaaaa gaaaaggggg 8761 gactggaagg gctaattcac tcccaacgaa gacaagatct gctttttgct tgtactgggt 8821 ctctctggtt agaccagatc tgagcctggg agctctctgg ctaactaggg aacccactgc 8881 ttaagcctca ataaagcttg ccttgagtgc ttcaagtagt gtgtgcccgt ctgttgtgtg 8941 actctggtaa ctagagatcc ctcagaccct tttagtcagt gtggaaaatc tctagcaggg 9001 cccgtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt 9061 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa 9121 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg 9181 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg 9241 gtgggctcta tggcttctga ggcggaaaga accagctggg gctctagggg gtatccccac 9301 gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct 9361 acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg 9421 ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt 9481 gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca 9541 tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga 9601 ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa 9661 gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac 9721 gcgaattaat tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag 9781 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc 9841 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag 9901 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc 9961 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc 10021 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg 10081 gagcttgtat atccattttc ggatctgatc agcacgtgtt gacaattaat catcggcata 10141 gtatatcggc atagtataat acgacaaggt gaggaactaa accatggcca agttgaccag 10201 tgccgttccg gtgctcaccg cgcgcgacgt cgccggagcg gtcgagttct ggaccgaccg 10261 gctcgggttc tcccgggact tcgtggagga cgacttcgcc ggtgtggtcc gggacgacgt 10321 gaccctgttc atcagcgcgg tccaggacca ggtggtgccg gacaacaccc tggcctgggt 10381 gtgggtgcgc ggcctggacg agctgtacgc cgagtggtcg gaggtcgtgt ccacgaactt 10441 ccgggacgcc tccgggccgg ccatgaccga gatcggcgag cagccgtggg ggcgggagtt 10501 cgccctgcgc gacccggccg gcaactgcgt gcacttcgtg gccgaggagc aggactgaca 10561 cgtgctacga gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt 10621 tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc 10681 ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa 10741 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa 10801 tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc 10861 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg 10921 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt 10981 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg 11041 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga 11101 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat 11161 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca 11221 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc 11281 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata 11341 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc 11401 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc 11461 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga 11521 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc 11581 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag 11641 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag 11701 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag 11761 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca 11821 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga 11881 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat 11941 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga 12001 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg 12061 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga 12121 gggcttacca tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc 12181 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac 12241 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc 12301 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc 12361 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc 12421 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt 12481 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc 12541 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg 12601 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag 12661 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat 12721 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc 12781 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa 12841 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta 12901 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa 12961 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg ac //