LOCUS Exported 13526 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION Lentiviral vector with puro selection for expression of S. aureus dCas9-KRAB. ACCESSION . VERSION . KEYWORDS pLV hUbC-dSaCas9-KRAB-T2A-PuroR SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 13526) AUTHORS Thakore PI, Kwon JB, Nelson CE, Rouse DC, Gemberling MP, Oliver ML, Gersbach CA TITLE RNA-guided transcriptional silencing in vivo with S. aureus CRISPR-Cas9 repressors. JOURNAL Nat Commun. 2018 Apr 26;9(1):1674. doi: 10.1038/s41467-018-04048-4. PUBMED 29700298 REFERENCE 2 (bases 1 to 13526) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..13526 /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 2602..3812 /label=UbC promoter /note="human ubiquitin C promoter" primer_bind 3649..3668 /label=hUBCpro-F /note="Human Ubiquitin C (UbC) promoter, forward primer" CDS 3850..3876 /codon_start=1 /product="HA (human influenza hemagglutinin) epitope tag" /label=HA /translation="YPYDVPDYA" CDS 3883..3903 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 7084..7131 /codon_start=1 /product="bipartite nuclear localization signal from nucleoplasmin" /label=nucleoplasmin NLS /translation="KRPAATKKAGQAKKKK" CDS 7165..7359 /codon_start=1 /product="Kruppel-associated box (KRAB) transcriptional repression domain from the human zinc finger protein ZNF10 (Margolin et al., 1994)" /label=KRAB /translation="RTLVTFKDVFVDFTREEWKLLDTAQQILYRNVMLENYKNLVSLGY QLTKPDVILRLEKGEEPWLV" CDS 7426..7446 /codon_start=1 /product="nuclear localization signal of SV40 (simian virus 40) large T antigen" /label=SV40 NLS /translation="PKKKRKV" CDS 7453..7506 /codon_start=1 /product="2A peptide from Thosea asigna virus capsid protein" /label=T2A /note="Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides." /translation="EGRGSLLTCGDVEENPGP" CDS 7507..8106 /codon_start=1 /gene="pac from Streptomyces alboniger" /product="puromycin N-acetyltransferase" /label=PuroR /note="confers resistance to puromycin" /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA" primer_bind complement(7507..7526) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 8003..8023 /label=Puro-F /note="Puromycin resistance gene, forward primer" protein_bind 8160..8193 /label=loxP /bound_moiety="Cre recombinase" /note="Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT)." misc_feature 8218..8806 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(8271..8291) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(8689..8700) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" primer_bind complement(8809..8825) /label=KS primer /note="common sequencing primer, one of multiple similar variants" primer_bind complement(8810..8826) /label=pBluescriptKS /note="For pBluescript vector" LTR 9331..9511 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" primer_bind complement(9537..9554) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 9543..9767 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 9813..10241 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(9900..9919) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 10110..10131 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(10250..10270) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 10255..10584 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 10435..10570 /label=SV40 ori /note="SV40 origin of replication" primer_bind 10497..10516 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" promoter 10632..10679 /label=EM7 promoter /note="synthetic bacterial promoter " CDS 10698..11072 /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 11202..11323 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(11239..11258) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 11293..11312 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(11372..11388) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(11372..11388) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(11385..11407) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 11396..11412 /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(11420..11450) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 11465..11486 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(11603..11620) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(11774..12362) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(11854..11873) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(12533..13393) /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 13156..13175 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(13394..13498) /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 aggcctccgc gccgggtttt ggcgcctccc gcgggcgccc 2641 ccctcctcac ggcgagcgct gccacgtcag acgaagggcg caggagcgtt cctgatcctt 2701 ccgcccggac gctcaggaca gcggcccgct gctcataaga ctcggcctta gaaccccagt 2761 atcagcagaa ggacatttta ggacgggact tgggtgactc tagggcactg gttttctttc 2821 cagagagcgg aacaggcgag gaaaagtagt cccttctcgg cgattctgcg gagggatctc 2881 cgtggggcgg tgaacgccga tgattatata aggacgcgcc gggtgtggca cagctagttc 2941 cgtcgcagcc gggatttggg tcgcggttct tgtttgtgga tcgctgtgat cgtcacttgg 3001 tgagttgcgg gctgctgggc tggccggggc tttcgtggcc gccgggccgc tcggtgggac 3061 ggaagcgtgt ggagagaccg ccaagggctg tagtctgggt ccgcgagcaa ggttgccctg 3121 aactgggggt tggggggagc gcacaaaatg gcggctgttc ccgagtcttg aatggaagac 3181 gcttgtaagg cgggctgtga ggtcgttgaa acaaggtggg gggcatggtg ggcggcaaga 3241 acccaaggtc ttgaggcctt cgctaatgcg ggaaagctct tattcgggtg agatgggctg 3301 gggcaccatc tggggaccct gacgtgaagt ttgtcactga ctggagaact cgggtttgtc 3361 gtctggttgc gggggcggca gttatgcggt gccgttgggc agtgcacccg tacctttggg 3421 agcgcgcgcc tcgtcgtgtc gtgacgtcac ccgttctgtt ggcttataat gcagggtggg 3481 gccacctgcc ggtaggtgtg cggtaggctt ttctccgtcg caggacgcag ggttcgggcc 3541 tagggtaggc tctcctgaat cgacaggcgc cggacctctg gtgaggggag ggataagtga 3601 ggcgtcagtt tctttggtcg gttttatgta cctatcttct taagtagctg aagctccggt 3661 tttgaactat gcgctcgggg ttggcgagtg tgttttgtga agttttttag gcaccttttg 3721 aaatgtaatc atttgggtca atatgtaatt ttcagtgtta gactagtaaa ttgtccgcta 3781 aattctggcc gtttttggct tttttgttag acgaagcttg ggctgcaggt cgactctaga 3841 gccaccatgt acccatacga tgttccagat tacgctatgg ccccaaagaa gaagcggaag 3901 gtcggtatcc acggagtccc agcagccaag cggaactaca tcctgggcct ggccatcggc 3961 atcaccagcg tgggctacgg catcatcgac tacgagacac gggacgtgat cgatgccggc 4021 gtgcggctgt tcaaagaggc caacgtggaa aacaacgagg gcaggcggag caagagaggc 4081 gccagaaggc tgaagcggcg gaggcggcat agaatccaga gagtgaagaa gctgctgttc 4141 gactacaacc tgctgaccga ccacagcgag ctgagcggca tcaaccccta cgaggccaga 4201 gtgaagggcc tgagccagaa gctgagcgag gaagagttct ctgccgccct gctgcacctg 4261 gccaagagaa gaggcgtgca caacgtgaac gaggtggaag aggacaccgg caacgagctg 4321 tccaccaaag agcagatcag ccggaacagc aaggccctgg aagagaaata cgtggccgaa 4381 ctgcagctgg aacggctgaa gaaagacggc gaagtgcggg gcagcatcaa cagattcaag 4441 accagcgact acgtgaaaga agccaaacag ctgctgaagg tgcagaaggc ctaccaccag 4501 ctggaccaga gcttcatcga cacctacatc gacctgctgg aaacccggcg gacctactat 4561 gagggacctg gcgagggcag ccccttcggc tggaaggaca tcaaagaatg gtacgagatg 4621 ctgatgggcc actgcaccta cttccccgag gaactgcgga gcgtgaagta cgcctacaac 4681 gccgacctgt acaacgccct gaacgacctg aacaatctcg tgatcaccag ggacgagaac 4741 gagaagctgg aatattacga gaagttccag atcatcgaga acgtgttcaa gcagaagaag 4801 aagcccaccc tgaagcagat cgccaaagaa atcctcgtga acgaagagga tattaagggc 4861 tacagagtga ccagcaccgg caagcccgag ttcaccaacc tgaaggtgta ccacgacatc 4921 aaggacatta ccgcccggaa agagattatt gagaacgccg agctgctgga tcagattgcc 4981 aagatcctga ccatctacca gagcagcgag gacatccagg aagaactgac caatctgaac 5041 tccgagctga cccaggaaga gatcgagcag atctctaatc tgaagggcta taccggcacc 5101 cacaacctga gcctgaaggc catcaacctg atcctggacg agctgtggca caccaacgac 5161 aaccagatcg ctatcttcaa ccggctgaag ctggtgccca agaaggtgga cctgtcccag 5221 cagaaagaga tccccaccac cctggtggac gacttcatcc tgagccccgt cgtgaagaga 5281 agcttcatcc agagcatcaa agtgatcaac gccatcatca agaagtacgg cctgcccaac 5341 gacatcatta tcgagctggc ccgcgagaag aactccaagg acgcccagaa aatgatcaac 5401 gagatgcaga agcggaaccg gcagaccaac gagcggatcg aggaaatcat ccggaccacc 5461 ggcaaagaga acgccaagta cctgatcgag aagatcaagc tgcacgacat gcaggaaggc 5521 aagtgcctgt acagcctgga agccatccct ctggaagatc tgctgaacaa ccccttcaac 5581 tatgaggtgg accacatcat ccccagaagc gtgtccttcg acaacagctt caacaacaag 5641 gtgctcgtga agcaggaaga agccagcaag aagggcaacc ggaccccatt ccagtacctg 5701 agcagcagcg acagcaagat cagctacgaa accttcaaga agcacatcct gaatctggcc 5761 aagggcaagg gcagaatcag caagaccaag aaagagtatc tgctggaaga acgggacatc 5821 aacaggttct ccgtgcagaa agacttcatc aaccggaacc tggtggatac cagatacgcc 5881 accagaggcc tgatgaacct gctgcggagc tacttcagag tgaacaacct ggacgtgaaa 5941 gtgaagtcca tcaatggcgg cttcaccagc tttctgcggc ggaagtggaa gtttaagaaa 6001 gagcggaaca aggggtacaa gcaccacgcc gaggacgccc tgatcattgc caacgccgat 6061 ttcatcttca aagagtggaa gaaactggac aaggccaaaa aagtgatgga aaaccagatg 6121 ttcgaggaaa agcaggccga gagcatgccc gagatcgaaa ccgagcagga gtacaaagag 6181 atcttcatca ccccccacca gatcaagcac attaaggact tcaaggacta caagtacagc 6241 caccgggtgg acaagaagcc taatagagag ctgattaacg acaccctgta ctccacccgg 6301 aaggacgaca agggcaacac cctgatcgtg aacaatctga acggcctgta cgacaaggac 6361 aatgacaagc tgaaaaagct gatcaacaag agccccgaaa agctgctgat gtaccaccac 6421 gacccccaga cctaccagaa actgaagctg attatggaac agtacggcga cgagaagaat 6481 cccctgtaca agtactacga ggaaaccggg aactacctga ccaagtactc caaaaaggac 6541 aacggccccg tgatcaagaa gattaagtat tacggcaaca aactgaacgc ccatctggac 6601 atcaccgacg actaccccaa cagcagaaac aaggtcgtga agctgtccct gaagccctac 6661 agattcgacg tgtacctgga caatggcgtg tacaagttcg tgaccgtgaa gaatctggat 6721 gtgatcaaaa aagaaaacta ctacgaagtg aatagcaagt gctatgagga agctaagaag 6781 ctgaagaaga tcagcaacca ggccgagttt atcgcctcct tctacaacaa cgatctgatc 6841 aagatcaacg gcgagctgta tagagtgatc ggcgtgaaca acgacctgct gaaccggatc 6901 gaagtgaaca tgatcgacat cacctaccgc gagtacctgg aaaacatgaa cgacaagagg 6961 ccccccagga tcattaagac aatcgcctcc aagacccaga gcattaagaa gtacagcaca 7021 gacattctgg gcaacctgta tgaagtgaaa tctaagaagc accctcagat catcaaaaag 7081 ggcaaaaggc cggcggccac gaaaaaggcc ggccaggcaa aaaagaaaaa gggatccgat 7141 gctaagtcac tgactgcctg gtcccggaca ctggtgacct tcaaggatgt gtttgtggac 7201 ttcaccaggg aggagtggaa gctgctggac actgctcagc agatcctgta cagaaatgtg 7261 atgctggaga actataagaa cctggtttcc ttgggttatc agcttactaa gccagatgtg 7321 atcctccggt tggagaaggg agaagagccc tggctggtgg agagagaaat tcaccaagag 7381 acccatcctg attcagagac tgcatttgaa atcaaatcat cagttccgaa aaagaaacgc 7441 aaagttgcta gcgagggcag aggaagtctt ctaacatgcg gtgacgtgga ggagaatccc 7501 ggccctatga ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga cgtccccagg 7561 gccgtacgca ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat 7621 ccggaccgcc acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac gcgcgtcggg 7681 ctcgacatcg gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt ctggaccacg 7741 ccggagagcg tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat ggccgagttg 7801 agcggttccc ggctggccgc gcagcaacag atggaaggcc tcctggcgcc gcaccggccc 7861 aaggagcccg cgtggttcct ggccaccgtc ggcgtgtcgc ccgaccacca gggcaagggt 7921 ctgggcagcg ccgtcgtgct ccccggagtg gaggcggccg agcgcgccgg ggtgcccgcc 7981 ttcctggaga cctccgcgcc ccgcaacctc cccttctacg agcggctcgg cttcaccgtc 8041 accgccgacg tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg caagcccggt 8101 gcctgaccag cacactggcg gccgttacta gcttctgcag cacgaccggt tgataataga 8161 taacttcgta tagcatacat tatacgaagt tatgaattcg atatcaagct tatcgataat 8221 caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct 8281 tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg 8341 gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg 8401 cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt 8461 tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt 8521 gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg 8581 ggcactgaca attccgtggt gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc 8641 tgtgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat 8701 ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc 8761 cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcatcg ataccgtcga 8821 cctcgagacc tagaaaaaca tggagcaatc acaagtagca atacagcagc taccaatgct 8881 gattgtgcct ggctagaagc acaagaggag gaggaggtgg gttttccagt cacacctcag 8941 gtacctttaa gaccaatgac ttacaaggca gctgtagatc ttagccactt tttaaaagaa 9001 aaggggggac tggaagggct aattcactcc caacgaagac aagatatcct tgatctgtgg 9061 atctaccaca cacaaggcta cttccctgat tggcagaact acacaccagg gccagggatc 9121 agatatccac tgacctttgg atggtgctac aagctagtac cagttgagca agagaaggta 9181 gaagaagcca atgaaggaga gaacacccgc ttgttacacc ctgtgagcct gcatgggatg 9241 gatgacccgg agagagaagt attagagtgg aggtttgaca gccgcctagc atttcatcac 9301 atggcccgag agctgcatcc ggactgtact gggtctctct ggttagacca gatctgagcc 9361 tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag cttgccttga 9421 gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag atccctcaga 9481 cccttttagt cagtgtggaa aatctctagc agggcccgtt taaacccgct gatcagcctc 9541 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac 9601 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg 9661 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga 9721 ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga 9781 aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc 9841 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc 9901 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct 9961 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa 10021 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc 10081 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact 10141 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg 10201 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt 10261 cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat 10321 ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg 10381 caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg 10441 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt 10501 tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt 10561 tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct 10621 gatcagcacg tgttgacaat taatcatcgg catagtatat cggcatagta taatacgaca 10681 aggtgaggaa ctaaaccatg gccaagttga ccagtgccgt tccggtgctc accgcgcgcg 10741 acgtcgccgg agcggtcgag ttctggaccg accggctcgg gttctcccgg gacttcgtgg 10801 aggacgactt cgccggtgtg gtccgggacg acgtgaccct gttcatcagc gcggtccagg 10861 accaggtggt gccggacaac accctggcct gggtgtgggt gcgcggcctg gacgagctgt 10921 acgccgagtg gtcggaggtc gtgtccacga acttccggga cgcctccggg ccggccatga 10981 ccgagatcgg cgagcagccg tgggggcggg agttcgccct gcgcgacccg gccggcaact 11041 gcgtgcactt cgtggccgag gagcaggact gacacgtgct acgagatttc gattccaccg 11101 ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc 11161 tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt 11221 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac 11281 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt 11341 cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt 11401 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg 11461 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg 11521 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc 11581 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc 11641 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata 11701 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg 11761 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct 11821 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa 11881 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc 11941 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt 12001 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg 12061 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg 12121 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct 12181 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc 12241 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg 12301 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc 12361 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt 12421 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa 12481 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat 12541 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct 12601 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg 12661 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag 12721 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta 12781 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg 12841 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg 12901 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct 12961 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta 13021 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg 13081 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc 13141 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg 13201 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga 13261 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg 13321 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat 13381 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc 13441 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca 13501 catttccccg aaaagtgcca cctgac //