LOCUS Exported 12352 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION 3rd generation lentiviral vector encoding FUCCI reporters for cell cycle analysis and puro for selection. ACCESSION . VERSION . KEYWORDS pBOB-EF1-FastFUCCI-Puro SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 12352) AUTHORS Koh SB, Mascalchi P, Rodriguez E, Lin Y, Jodrell DI, Richards FM, Lyons SK TITLE Quantitative FastFUCCI assay defines cell cycle dynamics at single-cell level. JOURNAL J Cell Sci. 2016 Nov 25. pii: jcs.195164. PUBMED 27888217 REFERENCE 2 (bases 1 to 12352) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..12352 /organism="synthetic DNA construct" /mol_type="other DNA" enhancer 172..551 /label=CMV enhancer /note="human cytomegalovirus immediate early enhancer" promoter 553..751 /label=CMV promoter /note="human cytomegalovirus (CMV) immediate early promoter" primer_bind 706..726 /label=CMV-F /note="Human CMV immediate early promoter, forward primer" LTR 769..949 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" misc_feature 996..1121 /label=HIV-1 Psi /note="packaging signal of human immunodeficiency virus type 1" misc_feature 1618..1851 /label=RRE /note="The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm." CDS 2036..2080 /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 2332..2448 /label=cPPT/CTS /note="central polypurine tract and central termination sequence of HIV-1" promoter 2465..3643 /label=EF-1-alpha promoter /note="strong constitutive promoter for human elongation factor EF-1-alpha" intron 2696..3634 /label=EF-1-alpha intron A /note="intron upstream of the start codon of human EF-1-alpha" primer_bind 3591..3611 /label=EF1a-F /note="Human elongation factor-1a promoter, forward primer" CDS 3664..4314 /codon_start=1 /product="monomeric Kusabira-Orange 2 fluorescent protein" /label=mKO2 /translation="MVSVIKPEMKMRYYMDGSVNGHEFTIEGEGTGRPYEGHQEMTLRV TMAEGGPMPFAFDLVSHVFCYGHRVFTKYPEEIPDYFKQAFPEGLSWERSLEFEDGGSA SVSAHISLRGNTFYHKSKFTGVNFPADGPIMQNQSVDWEPSTEKITASDGVLKGDVTMY LKLEGGGNHKCQMKTTYKAAKEILEMPGDHYIGHRLVRKTEGNITEQVEDAVAH" CDS 4369..4641 /codon_start=1 /product="degron consisting of residues 30-120 of human Cdt1 (Zielke and Edgar, 2015)" /label=Cdt1 (30-120) /translation="PSPARPALRAPASATSGSRKRARPPAAPGRDQARPPARRRLRLSV DEVSSPSTPEAPDIPACPSPGQKIKKSTPAAGQPPHLTSAQDQDTI" CDS 4648..4701 /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 4708..5385 /codon_start=1 /product="monomeric Azami-Green fluorescent protein (Karasawa et al., 2003)" /label=mAzami-Green /translation="MVSVIKPEMKIKLCMRGTVNGHNFVIEGEGKGNPYEGTQILDLNV TEGAPLPFAYDILTTVFQYGNRAFTKYPADIQDYFKQTFPEGYHWERSMTYEDQGICTA TSNISMRGDCFFYDIRFDGTNFPPNGPVMQKKTLKWEPSTEKMYVEDGVLKGDVNMRLL LEGGGHYRCDFKTTYKAKKEVRLPDAHKIDHRIEILKHDKDYNKVKLYENAVARYSMLP SQAK" CDS 5416..5745 /codon_start=1 /product="degron consisting of residues 1-110 of human Geminin (Zielke and Edgar, 2015)" /label=Gem1 (1-110) /translation="MNPSMKQKQEEIKENIKNSSVPRRTLKMIQPSASGSLVGRENELS AGLSKRKHRNDHLTSTTSSPGVIVPESSENKNLGGVTQESFDLMIKENPSSQYWKEVAE KRRKAL" promoter 5788..6287 /label=PGK promoter /note="mouse phosphoglycerate kinase 1 promoter" primer_bind complement(5823..5846) /label=MSCV-rev /note="Murine stem cell virus, reverse primer" primer_bind 6220..6239 /label=mPGK-F /note="Mouse PGK promoter, forward primer" CDS 6331..6930 /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(6331..6350) /label=Puro-R /note="Puromycin resistance gene, reverse primer. Also called puro-variant-R" primer_bind 6827..6847 /label=Puro-F /note="Puromycin resistance gene, forward primer" misc_feature 7092..7680 /label=WPRE /note="woodchuck hepatitis virus posttranscriptional regulatory element" primer_bind complement(7145..7165) /label=WPRE-R /note="WPRE, reverse primer" CDS complement(7563..7574) /codon_start=1 /product="Factor Xa recognition and cleavage site" /label=Factor Xa site /translation="IEGR" LTR 8091..8271 /label=5' LTR (truncated) /note="truncated 5' long terminal repeat (LTR) from HIV-1" primer_bind complement(8297..8314) /label=BGH-rev /note="Bovine growth hormone terminator, reverse primer. Also called BGH reverse" polyA_signal 8303..8527 /label=bGH poly(A) signal /note="bovine growth hormone polyadenylation signal" rep_origin 8573..9001 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(8660..8679) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 8870..8891 /label=F1ori-F /note="F1 origin, forward primer" primer_bind complement(9010..9030) /label=pBABE 3' /note="SV40 enhancer, reverse primer for pBABE vectors" promoter 9015..9344 /label=SV40 promoter /note="SV40 enhancer and early promoter" rep_origin 9195..9330 /label=SV40 ori /note="SV40 origin of replication" primer_bind 9257..9276 /label=SV40pro-F /note="SV40 promoter/origin, forward primer" promoter 9392..9439 /label=EM7 promoter /note="synthetic bacterial promoter " CDS 9458..9832 /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 9962..10083 /label=SV40 poly(A) signal /note="SV40 polyadenylation signal" primer_bind complement(9999..10018) /label=SV40pA-R /note="SV40 polyA, reverse primer" primer_bind 10053..10072 /label=EBV-rev /note="SV40 polyA terminator, reverse primer" primer_bind complement(10132..10148) /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind complement(10132..10148) /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" primer_bind complement(10145..10167) /label=M13/pUC Reverse /note="In lacZ gene" protein_bind 10156..10172 /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(10180..10210) /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 10225..10246 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." primer_bind complement(10363..10380) /label=L4440 /note="L4440 vector, forward primer" rep_origin complement(10534..11122) /direction=LEFT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind complement(10614..10633) /label=pBR322ori-F /note="pBR322 origin, forward primer" CDS complement(11293..12153) /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 11916..11935 /label=Amp-R /note="Ampicillin resistance gene, reverse primer" promoter complement(12154..12258) /gene="bla" /label=AmpR promoter primer_bind complement(12333..12352) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" ORIGIN 1 catagttaag ccagtatctg ctccctgctt gtgtgttgga ggtcgctgag tagtgcgcga 61 gcaaaattta agctacaaca aggcaaggct tgaccgacaa ttgcatgaag aatctgctta 121 gggttaggcg ttttgcgctg cttcgcgatg tacgggccag atatacgcgt tgacattgat 181 tattgactag ttattaatag taatcaatta cggggtcatt agttcatagc ccatatatgg 241 agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc 301 gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg actttccatt 361 gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat caagtgtatc 421 atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc tggcattatg 481 cccagtacat gaccttatgg gactttccta cttggcagta catctacgta ttagtcatcg 541 ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact 601 cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa 661 atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta 721 ggcgtgtacg gtgggaggtc tatataagca gcgcgttttg cctgtactgg gtctctctgg 781 ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct 841 caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt 901 aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag tggcgcccga 961 acagggactt gaaagcgaaa gggaaaccag aggagctctc tcgacgcagg actcggcttg 1021 ctgaagcgcg cacggcaaga ggcgaggggc ggcgactggt gagtacgcca aaaattttga 1081 ctagcggagg ctagaaggag agagatgggt gcgagagcgt cagtattaag cgggggagaa 1141 ttagatcgcg atgggaaaaa attcggttaa ggccaggggg aaagaaaaaa tataaattaa 1201 aacatatagt atgggcaagc agggagctag aacgattcgc agttaatcct ggcctgttag 1261 aaacatcaga aggctgtaga caaatactgg gacagctaca accatccctt cagacaggat 1321 cagaagaact tagatcatta tataatacag tagcaaccct ctattgtgtg catcaaagga 1381 tagagataaa agacaccaag gaagctttag acaagataga ggaagagcaa aacaaaagta 1441 agaccaccgc acagcaagcg gccggccgct gatcttcaga cctggaggag gagatatgag 1501 ggacaattgg agaagtgaat tatataaata taaagtagta aaaattgaac cattaggagt 1561 agcacccacc aaggcaaaga gaagagtggt gcagagagaa aaaagagcag tgggaatagg 1621 agctttgttc cttgggttct tgggagcagc aggaagcact atgggcgcag cgtcaatgac 1681 gctgacggta caggccagac aattattgtc tggtatagtg cagcagcaga acaatttgct 1741 gagggctatt gaggcgcaac agcatctgtt gcaactcaca gtctggggca tcaagcagct 1801 ccaggcaaga atcctggctg tggaaagata cctaaaggat caacagctcc tggggatttg 1861 gggttgctct ggaaaactca tttgcaccac tgctgtgcct tggaatgcta gttggagtaa 1921 taaatctctg gaacagattt ggaatcacac gacctggatg gagtgggaca gagaaattaa 1981 caattacaca agcttaatac actccttaat tgaagaatcg caaaaccagc aagaaaagaa 2041 tgaacaagaa ttattggaat tagataaatg ggcaagtttg tggaattggt ttaacataac 2101 aaattggctg tggtatataa aattattcat aatgatagta ggaggcttgg taggtttaag 2161 aatagttttt gctgtacttt ctatagtgaa tagagttagg cagggatatt caccattatc 2221 gtttcagacc cacctcccaa ccccgagggg acccgacagg cccgaaggaa tagaagaaga 2281 aggtggagag agagacagag acagatccat tcgattagtg aacggatcaa cttttaaaag 2341 aaaagggggg attggggggt acagtgcagg ggaaagaata gtagacataa tagcaacaga 2401 catacaaact aaagaattac aaaaacaaat tacaaaattc aaaattttat cttaattaag 2461 aattggctcc ggtgcccgtc agtgggcaga gcgcacatcg cccacagtcc ccgagaagtt 2521 ggggggaggg gtcggcaatt gaaccggtgc ctagagaagg tggcgcgggg taaactggga 2581 aagtgatgtc gtgtactggc tccgcctttt tcccgagggt gggggagaac cgtatataag 2641 tgcagtagtc gccgtgaacg ttctttttcg caacgggttt gccgccagaa cacaggtaag 2701 tgccgtgtgt ggttcccgcg ggcctggcct ctttacgggt tatggccctt gcgtgccttg 2761 aattacttcc acctggctgc agtacgtgat tcttgatccc gagcttcggg ttggaagtgg 2821 gtgggagagt tcgaggcctt gcgcttaagg agccccttcg cctcgtgctt gagttgaggc 2881 ctggcctggg cgctggggcc gccgcgtgcg aatctggtgg caccttcgcg cctgtctcgc 2941 tgctttcgat aagtctctag ccatttaaaa tttttgatga cctgctgcga cgcttttttt 3001 ctggcaagat agtcttgtaa atgcgggcca agatctgcac actggtattt cggtttttgg 3061 ggccgcgggc ggcgacgggg cccgtgcgtc ccagcgcaca tgttcggcga ggcggggcct 3121 gcgagcgcgg ccaccgagaa tcggacgggg gtagtctcaa gctggccggc ctgctctggt 3181 gcctggcctc gcgccgccgt gtatcgcccc gccctgggcg gcaaggctgg cccggtcggc 3241 accagttgcg tgagcggaaa gatggccgct tcccggccct gctgcaggga gctcaaaatg 3301 gaggacgcgg cgctcgggag agcgggcggg tgagtcaccc acacaaagga aaagggcctt 3361 tccgtcctca gccgtcgctt catgtgactc cacggagtac cgggcgccgt ccaggcacct 3421 cgattagttc tcgagctttt ggagtacgtc gtctttaggt tggggggagg ggttttatgc 3481 gatggagttt ccccacactg agtgggtgga gactgaagtt aggccagctt ggcacttgat 3541 gtaattctcc ttggaatttg ccctttttga gtttggatct tggttcattc tcaagcctca 3601 gacagtggtt caaagttttt ttcttccatt tcaggtgtcg tgattaatta agaattggga 3661 tccatggtgt ccgtgatcaa gcccgagatg aagatgcggt actacatgga cggcagcgtg 3721 aacggccacg agttcaccat cgagggcgag ggcaccggca gaccttacga gggccaccag 3781 gaaatgaccc tgagagtcac catggccgag ggcggaccca tgcccttcgc cttcgacctg 3841 gtgtcccacg tgttctgcta cggccaccgg gtgttcacca agtaccccga ggaaatcccc 3901 gactacttca agcaggcctt ccccgagggc ctgagctggg agcggagcct ggaattcgag 3961 gatggcggca gcgcctctgt gtctgcccac atcagcctgc ggggcaacac cttctaccac 4021 aagagcaagt tcaccggcgt gaacttcccc gccgacggcc ccatcatgca gaaccagagc 4081 gtggactggg agcccagcac cgagaagatc accgccagcg acggcgtgct gaagggcgac 4141 gtgaccatgt acctgaagct ggaaggcggc ggaaaccaca agtgccagat gaaaaccacc 4201 tacaaggccg ccaaagaaat cctggaaatg cccggcgacc actacatcgg ccaccggctg 4261 gtgcggaaaa ccgagggcaa catcaccgag caggtggagg atgccgtggc ccactacagc 4321 atgctgccta gcctcggcgg ctaccctagc cattggaggc ccctggaacc ctctcctgcc 4381 agacctgccc tgagagcccc tgcctccgcc acctccggca gcagaaagag agccagacct 4441 cctgccgccc ctggcagaga tcaggccagg ccccctgccc gcagacggct ccggctgtcc 4501 gtggatgagg tgtccagccc tagcaccccc gaggcccccg atatccctgc ctgtcccagc 4561 cctggccaga aaatcaagaa gagcacacct gccgccggac agcctcctca cctgaccagc 4621 gcccaggacc aggacaccat cgtcgacgag ggcaggggca gcctgctgac ctgcggcgac 4681 gtggaggaga accccggccc cagatccatg gtgtccgtga tcaagcccga gatgaagatc 4741 aagctgtgca tgcggggaac cgtgaacggc cacaacttcg tgatcgaggg cgagggcaag 4801 ggaaacccct acgagggcac ccagatcctg gacctgaacg tgaccgaagg cgcccctctg 4861 cccttcgcct acgacatcct gaccaccgtg ttccagtacg gcaaccgggc cttcaccaag 4921 taccccgccg acatccagga ctacttcaag cagaccttcc ccgagggcta ccactgggag 4981 cggagcatga cctacgagga ccagggcatc tgcaccgcca cctccaacat cagcatgagg 5041 ggcgactgct ttttctacga catcagattc gacggcacca acttcccccc caacggcccc 5101 gtgatgcaga aaaagaccct gaagtgggag cccagcaccg agaagatgta cgtggaggac 5161 ggcgtgctga agggcgacgt gaacatgcgg ctgctgctgg aaggcggcgg acactacaga 5221 tgcgacttca agaccaccta caaggccaag aaagaagtgc ggctgcccga cgcccacaag 5281 atcgaccacc ggatcgagat cctgaagcac gacaaggact acaacaaagt gaagctgtac 5341 gagaacgccg tggccaggta cagcatgctg cccagccagg ccaagggcta ccccagccac 5401 tggcggcctc tggaaatgaa ccccagcatg aagcagaagc aggaagagat caaagagaac 5461 atcaagaaca gcagcgtgcc caggcggacc ctgaagatga tccagcccag cgccagcgga 5521 tctctggtgg gccgggagaa tgagctgtct gccggcctga gcaagcggaa gcaccggaac 5581 gaccacctga ccagcaccac ctctagccca ggcgtgatcg tgcccgagag cagcgagaac 5641 aagaacctgg gcggcgtgac ccaagaaagc ttcgacctga tgatcaaaga aaaccccagc 5701 agccagtact ggaaagaggt ggccgagaag cggcggaagg ccctgtaggt cgagcaattc 5761 gcggccgcgc tagctcgaaa ttctaccggg taggggaggc gcttttccca aggcagtctg 5821 gagcatgcgc tttagcagcc ccgctgggca cttggcgcta cacaagtggc ctctggcctc 5881 gcacacattc cacatccacc ggtaggcgcc aaccggctcc gttctttggt ggccccttcg 5941 cgccaccttc tactcctccc ctagtcagga agttcccccc cgccccgcag ctcgcgtcgt 6001 gcaggacgtg acaaatggaa gtagcacgtc tcactagtct cgtgcagatg gacagcaccg 6061 ctgagcaatg gaagcgggta ggcctttggg gcagcggcca atagcagctt tgctccttcg 6121 ctttctgggc tcagaggctg ggaaggggtg ggtccggggg cgggctcagg ggcgggctca 6181 ggggcggggc gggcgcccga aggtcctccg gaggcccggc attctgcacg cttcaaaagc 6241 gcacgtctgc cgcgctgttc tcctcttcct catctccggg cctttcgacc tgcatccatc 6301 tagatctcga tcgagcagct gaagcttacc atgaccgagt acaagcccac ggtgcgcctc 6361 gccacccgcg acgacgtccc cagggccgta cgcaccctcg ccgccgcgtt cgccgactac 6421 cccgccacgc gccacaccgt cgatccggac cgccacatcg agcgggtcac cgagctgcaa 6481 gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg tgtgggtcgc ggacgacggc 6541 gccgcggtgg cggtctggac cacgccggag agcgtcgaag cgggggcggt gttcgccgag 6601 atcggcccgc gcatggccga gttgagcggt tcccggctgg ccgcgcagca acagatggaa 6661 ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt tcctggccac cgtcggcgtc 6721 tcgcccgacc accagggcaa gggtctgggc agcgccgtcg tgctccccgg agtggaggcg 6781 gccgagcgcg ccggggtgcc cgccttcctg gagacctccg cgccccgcaa cctccccttc 6841 tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg tgcccgaagg accgcgcacc 6901 tggtgcatga cccgcaagcc cggtgcctga gctagctgca ggtcgagcat gcatctaggg 6961 cggccaattc cgccgatcct ggaaaaacat ggagcaatca caagtagcaa tacagcagct 7021 accaatgctg attgtgcctg gctagaagca caagaggagg aggaggtggg ttttccagtc 7081 acacctcaga caatcaacct ctggattaca aaatttgtga aagattgact ggtattctta 7141 actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg tatcatgcta 7201 ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggttg ctgtctcttt 7261 atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg 7321 caacccccac tggttggggc attgccacca cctgtcagct cctttccggg actttcgctt 7381 tccccctccc tattgccacg gcggaactca tcgccgcctg ccttgcccgc tgctggacag 7441 gggctcggct gttgggcact gacaattccg tggtgttgtc ggggaagctg acgtcctttc 7501 catggctgct cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc 7561 cttcggccct caatccagcg gaccttcctt cccgcggcct gctgccggct ctgcggcctc 7621 ttccgcgtct tcgccttcgc cctcagacga gtcggatctc cctttgggcc gcctccccgc 7681 ctggaattaa ttcgagctcg gtacctttaa gaccaatgac ttacaaggca gctgtagatc 7741 ttagccactt tttaaaagaa aaggggggac tggaagggct aattcactcc caacgaagac 7801 aagatatcct tgatctgtgg atctaccaca cacaaggcta cttccctgat tggcagaact 7861 acacaccagg gccagggatc agatatccac tgacctttgg atggtgctac aagctagtac 7921 cagttgagca agagaaggta gaagaagcca atgaaggaga gaacacccgc ttgttacacc 7981 ctgtgagcct gcatgggatg gatgacccgg agagagaagt attagagtgg aggtttgaca 8041 gccgcctagc atttcatcac atggcccgag agctgcatcc ggactgtact gggtctctct 8101 ggttagacca gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc 8161 ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg 8221 gtaactagag atccctcaga cccttttagt cagtgtggaa aatctctagc agggccgctt 8281 taaacccgct gatcagcctc gactgtgcct tctagttgcc agccatctgt tgtttgcccc 8341 tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc ctaataaaat 8401 gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg tggggtgggg 8461 caggacagca agggggagga ttgggaagac aatagcaggc atgctgggga tgcggtgggc 8521 tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc 8581 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt 8641 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc 8701 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta 8761 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtagtgg gccatcgccc 8821 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg 8881 ttccaaactg gaacaacact caaccctatc tcggtctatt cttttgattt ataagggatt 8941 ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat 9001 taattctgtg gaatgtgtgt cagttagggt gtggaaagtc cccaggctcc ccagcaggca 9061 gaagtatgca aagcatgcat ctcaattagt cagcaaccag gtgtggaaag tccccaggct 9121 ccccagcagg cagaagtatg caaagcatgc atctcaatta gtcagcaacc atagtcccgc 9181 ccctaactcc gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg 9241 gctgactaat tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc 9301 agaagtagtg aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt 9361 gtatatccat tttcggatct gatcagcacg tgttgacaat taatcatcgg catagtatat 9421 cggcatagta taatacgaca aggtgaggaa ctaaaccatg gccaagttga ccagtgccgt 9481 tccggtgctc accgcgcgcg acgtcgccgg agcggtcgag ttctggaccg accggctcgg 9541 gttctcccgg gacttcgtgg aggacgactt cgccggtgtg gtccgggacg acgtgaccct 9601 gttcatcagc gcggtccagg accaggtggt gccggacaac accctggcct gggtgtgggt 9661 gcgcggcctg gacgagctgt acgccgagtg gtcggaggtc gtgtccacga acttccggga 9721 cgcctccggg ccggccatga ccgagatcgg cgagcagccg tgggggcggg agttcgccct 9781 gcgcgacccg gccggcaact gcgtgcactt cgtggccgag gagcaggact gacacgtgct 9841 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg 9901 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc 9961 caacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac 10021 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc 10081 ttatcatgtc tgtataccgt cgacctctag ctagagcttg gcgtaatcat ggtcatagct 10141 gtttcctgtg tgaaattgtt atccgctcac aattccacac aacatacgag ccggaagcat 10201 aaagtgtaaa gcctggggtg cctaatgagt gagctaactc acattaattg cgttgcgctc 10261 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg 10321 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct 10381 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt 10441 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc 10501 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga 10561 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata 10621 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac 10681 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg 10741 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc 10801 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag 10861 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt 10921 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt 10981 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg 11041 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac 11101 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca 11161 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac 11221 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac 11281 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt 11341 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt 11401 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt 11461 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc 11521 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa 11581 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg 11641 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt 11701 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc 11761 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt 11821 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg 11881 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac 11941 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc 12001 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt 12061 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg 12121 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag 12181 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa 12241 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg acggatcggg 12301 agatctcccg atcccctatg gtgcactctc agtacaatct gctctgatgc cg //