LentiCRISPRV2GFP哺乳编辑质粒

基本信息

原核抗性:	氨苄青霉素Amp
筛选标记:	博来霉素Zeo，绿色荧光GFP
克隆菌株:	大肠杆菌Stbl3
培养条件:	37℃

质粒简介

           pLentiCRISPRV2GFP是一个哺乳细胞Cas9和gRNA表达质粒。

质粒图谱

质粒序列

LOCUS       Exported               13131 bp ds-DNA     circular SYN 07-JUN-2018
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    LentiCRISPRv2GFP
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13131)
  AUTHORS   Triple Threat
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..13131
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        1..241
                     /note="U6 promoter"
                     /note="RNA polymerase III promoter for human U6 snRNA"
     misc_RNA        266..341
                     /note="gRNA scaffold"
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        403..614
                     /note="EF-1-alpha core promoter"
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             639..4742
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /note="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             4743..4790
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /note="nucleoplasmin NLS"
                     /translation="KRPAATKKAGQAKKKK"
     CDS             4791..4814
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /note="FLAG"
                     /translation="DYKDDDDK"
     CDS             4824..4880
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /note="P2A"
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     misc_feature    5616..6204
                     /note="WPRE"
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     LTR             6276..6509
                     /note="3' LTR (Delta-U3)"
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    6541..6765
                     /note="bGH poly(A) signal"
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      6811..7239
                     /direction=RIGHT
                     /note="f1 ori"
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        7253..7582
                     /note="SV40 promoter"
                     /note="SV40 enhancer and early promoter"
     rep_origin      7433..7568
                     /note="SV40 ori"
                     /note="SV40 origin of replication"
     promoter        7630..7677
                     /note="EM7 promoter"
                     /note="synthetic bacterial promoter "
     CDS             7696..8070
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /note="Zeo"
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    8200..8321
                     /note="SV40 poly(A) signal"
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(8772..9360)
                     /direction=LEFT
                     /note="ori"
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(9531..10391)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /note="AmpR"
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(10392..10496)
                     /gene="bla"
                     /note="AmpR promoter"
     enhancer        10762..11141
                     /note="CMV enhancer"
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        11143..11341
                     /note="CMV promoter"
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     LTR             11359..11539
                     /note="5' LTR (truncated)"
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    11586..11711
                     /note="HIV-1 Psi"
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    12204..12437
                     /note="RRE"
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     misc_feature    12964..13081
                     /note="cPPT/CTS"
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
       61 ataattagaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
      121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
      181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
      241 cgaaacaccg gagacggacg tctctgtttt agagctagaa atagcaagtt aaaataaggc
      301 tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg cttttttgaa ttcgctagct
      361 aggtcttgaa aggagtggga attggctccg gtgcccgtca gtgggcagag cgcacatcgc
      421 ccacagtccc cgagaagttg gggggagggg tcggcaattg atccggtgcc tagagaaggt
      481 ggcgcggggt aaactgggaa agtgatgtcg tgtactggct ccgccttttt cccgagggtg
      541 ggggagaacc gtatataagt gcagtagtcg ccgtgaacgt tctttttcgc aacgggtttg
      601 ccgccagaac acaggaccgg ttctagagcg ctgccaccat ggacaagaag tacagcatcg
      661 gcctggacat cggcaccaac tctgtgggct gggccgtgat caccgacgag tacaaggtgc
      721 ccagcaagaa attcaaggtg ctgggcaaca ccgaccggca cagcatcaag aagaacctga
      781 tcggagccct gctgttcgac agcggcgaaa cagccgaggc cacccggctg aagagaaccg
      841 ccagaagaag atacaccaga cggaagaacc ggatctgcta tctgcaagag atcttcagca
      901 acgagatggc caaggtggac gacagcttct tccacagact ggaagagtcc ttcctggtgg
      961 aagaggataa gaagcacgag cggcacccca tcttcggcaa catcgtggac gaggtggcct
     1021 accacgagaa gtaccccacc atctaccacc tgagaaagaa actggtggac agcaccgaca
     1081 aggccgacct gcggctgatc tatctggccc tggcccacat gatcaagttc cggggccact
     1141 tcctgatcga gggcgacctg aaccccgaca acagcgacgt ggacaagctg ttcatccagc
     1201 tggtgcagac ctacaaccag ctgttcgagg aaaaccccat caacgccagc ggcgtggacg
     1261 ccaaggccat cctgtctgcc agactgagca agagcagacg gctggaaaat ctgatcgccc
     1321 agctgcccgg cgagaagaag aatggcctgt tcggaaacct gattgccctg agcctgggcc
     1381 tgacccccaa cttcaagagc aacttcgacc tggccgagga tgccaaactg cagctgagca
     1441 aggacaccta cgacgacgac ctggacaacc tgctggccca gatcggcgac cagtacgccg
     1501 acctgtttct ggccgccaag aacctgtccg acgccatcct gctgagcgac atcctgagag
     1561 tgaacaccga gatcaccaag gcccccctga gcgcctctat gatcaagaga tacgacgagc
     1621 accaccagga cctgaccctg ctgaaagctc tcgtgcggca gcagctgcct gagaagtaca
     1681 aagagatttt cttcgaccag agcaagaacg gctacgccgg ctacattgac ggcggagcca
     1741 gccaggaaga gttctacaag ttcatcaagc ccatcctgga aaagatggac ggcaccgagg
     1801 aactgctcgt gaagctgaac agagaggacc tgctgcggaa gcagcggacc ttcgacaacg
     1861 gcagcatccc ccaccagatc cacctgggag agctgcacgc cattctgcgg cggcaggaag
     1921 atttttaccc attcctgaag gacaaccggg aaaagatcga gaagatcctg accttccgca
     1981 tcccctacta cgtgggccct ctggccaggg gaaacagcag attcgcctgg atgaccagaa
     2041 agagcgagga aaccatcacc ccctggaact tcgaggaagt ggtggacaag ggcgcttccg
     2101 cccagagctt catcgagcgg atgaccaact tcgataagaa cctgcccaac gagaaggtgc
     2161 tgcccaagca cagcctgctg tacgagtact tcaccgtgta taacgagctg accaaagtga
     2221 aatacgtgac cgagggaatg agaaagcccg ccttcctgag cggcgagcag aaaaaggcca
     2281 tcgtggacct gctgttcaag accaaccgga aagtgaccgt gaagcagctg aaagaggact
     2341 acttcaagaa aatcgagtgc ttcgactccg tggaaatctc cggcgtggaa gatcggttca
     2401 acgcctccct gggcacatac cacgatctgc tgaaaattat caaggacaag gacttcctgg
     2461 acaatgagga aaacgaggac attctggaag atatcgtgct gaccctgaca ctgtttgagg
     2521 acagagagat gatcgaggaa cggctgaaaa cctatgccca cctgttcgac gacaaagtga
     2581 tgaagcagct gaagcggcgg agatacaccg gctggggcag gctgagccgg aagctgatca
     2641 acggcatccg ggacaagcag tccggcaaga caatcctgga tttcctgaag tccgacggct
     2701 tcgccaacag aaacttcatg cagctgatcc acgacgacag cctgaccttt aaagaggaca
     2761 tccagaaagc ccaggtgtcc ggccagggcg atagcctgca cgagcacatt gccaatctgg
     2821 ccggcagccc cgccattaag aagggcatcc tgcagacagt gaaggtggtg gacgagctcg
     2881 tgaaagtgat gggccggcac aagcccgaga acatcgtgat cgaaatggcc agagagaacc
     2941 agaccaccca gaagggacag aagaacagcc gcgagagaat gaagcggatc gaagagggca
     3001 tcaaagagct gggcagccag atcctgaaag aacaccccgt ggaaaacacc cagctgcaga
     3061 acgagaagct gtacctgtac tacctgcaga atgggcggga tatgtacgtg gaccaggaac
     3121 tggacatcaa ccggctgtcc gactacgatg tggaccatat cgtgcctcag agctttctga
     3181 aggacgactc catcgacaac aaggtgctga ccagaagcga caagaaccgg ggcaagagcg
     3241 acaacgtgcc ctccgaagag gtcgtgaaga agatgaagaa ctactggcgg cagctgctga
     3301 acgccaagct gattacccag agaaagttcg acaatctgac caaggccgag agaggcggcc
     3361 tgagcgaact ggataaggcc ggcttcatca agagacagct ggtggaaacc cggcagatca
     3421 caaagcacgt ggcacagatc ctggactccc ggatgaacac taagtacgac gagaatgaca
     3481 agctgatccg ggaagtgaaa gtgatcaccc tgaagtccaa gctggtgtcc gatttccgga
     3541 aggatttcca gttttacaaa gtgcgcgaga tcaacaacta ccaccacgcc cacgacgcct
     3601 acctgaacgc cgtcgtggga accgccctga tcaaaaagta ccctaagctg gaaagcgagt
     3661 tcgtgtacgg cgactacaag gtgtacgacg tgcggaagat gatcgccaag agcgagcagg
     3721 aaatcggcaa ggctaccgcc aagtacttct tctacagcaa catcatgaac tttttcaaga
     3781 ccgagattac cctggccaac ggcgagatcc ggaagcggcc tctgatcgag acaaacggcg
     3841 aaaccgggga gatcgtgtgg gataagggcc gggattttgc caccgtgcgg aaagtgctga
     3901 gcatgcccca agtgaatatc gtgaaaaaga ccgaggtgca gacaggcggc ttcagcaaag
     3961 agtctatcct gcccaagagg aacagcgata agctgatcgc cagaaagaag gactgggacc
     4021 ctaagaagta cggcggcttc gacagcccca ccgtggccta ttctgtgctg gtggtggcca
     4081 aagtggaaaa gggcaagtcc aagaaactga agagtgtgaa agagctgctg gggatcacca
     4141 tcatggaaag aagcagcttc gagaagaatc ccatcgactt tctggaagcc aagggctaca
     4201 aagaagtgaa aaaggacctg atcatcaagc tgcctaagta ctccctgttc gagctggaaa
     4261 acggccggaa gagaatgctg gcctctgccg gcgaactgca gaagggaaac gaactggccc
     4321 tgccctccaa atatgtgaac ttcctgtacc tggccagcca ctatgagaag ctgaagggct
     4381 cccccgagga taatgagcag aaacagctgt ttgtggaaca gcacaagcac tacctggacg
     4441 agatcatcga gcagatcagc gagttctcca agagagtgat cctggccgac gctaatctgg
     4501 acaaagtgct gtccgcctac aacaagcacc gggataagcc catcagagag caggccgaga
     4561 atatcatcca cctgtttacc ctgaccaatc tgggagcccc tgccgccttc aagtactttg
     4621 acaccaccat cgaccggaag aggtacacca gcaccaaaga ggtgctggac gccaccctga
     4681 tccaccagag catcaccggc ctgtacgaga cacggatcga cctgtctcag ctgggaggcg
     4741 acaagcgacc tgccgccaca aagaaggctg gacaggctaa gaagaagaaa gattacaaag
     4801 acgatgacga taagggatcc ggcgcaacaa acttctctct gctgaaacaa gccggagatg
     4861 tcgaagagaa tcctggaccg atggtgtcta agggcgaaga gctgttcacc ggggtggtgc
     4921 ccatcctggt cgagctggac ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg
     4981 gcgagggcga tgccacctac ggcaagctga ccctgaagtt catctgcacc accggcaagc
     5041 tgcccgtgcc ctggcccacc ctcgtgacca ccctgaccta cggcgtgcag tgcttcagcc
     5101 gctaccccga ccacatgaag cagcacgact tcttcaagtc cgccatgccc gaaggctacg
     5161 tccaggagcg caccatcttc ttcaaggacg acggcaacta caagacccgc gccgaggtga
     5221 agttcgaggg cgacaccctg gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg
     5281 acggcaacat cctggggcac aagctggagt acaactacaa cagccacaac gtctatatca
     5341 tggccgacaa gcagaagaac ggcatcaagg tgaacttcaa gatccgccac aacatcgagg
     5401 acggcagcgt gcagctcgcc gaccactacc agcagaacac ccccatcggc gacggccccg
     5461 tgctgctgcc cgacaaccac tacctgagca cccagtccgc cctgagcaaa gaccccaacg
     5521 agaagcgcga tcacatggtc ctgctggagt tcgtgaccgc cgccgggatc actctcggca
     5581 tggacgagct gtacaagtaa acgcgttaag tcgacaatca acctctggat tacaaaattt
     5641 gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg
     5701 ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt
     5761 ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg
     5821 tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc
     5881 agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg
     5941 cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt
     6001 tgtcggggaa atcatcgtcc tttccttggc tgctcgcctg tgttgccacc tggattctgc
     6061 gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg
     6121 gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga
     6181 tctccctttg ggccgcctcc ccgcgtcgac tttaagacca atgacttaca aggcagctgt
     6241 agatcttagc cactttttaa aagaaaaggg gggactggaa gggctaattc actcccaacg
     6301 aagacaagat ctgctttttg cttgtactgg gtctctctgg ttagaccaga tctgagcctg
     6361 ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct tgccttgagt
     6421 gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat ccctcagacc
     6481 cttttagtca gtgtggaaaa tctctagcag ggcccgttta aacccgctga tcagcctcga
     6541 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     6601 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     6661 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     6721 gggaagacaa tagcaggcat gctggggatg cggtgggctc tatggcttct gaggcggaaa
     6781 gaaccagctg gggctctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg
     6841 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     6901 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     6961 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     7021 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     7081 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     7141 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     7201 taaaaaatga gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca
     7261 gttagggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa gcatgcatct
     7321 caattagtca gcaaccaggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca
     7381 aagcatgcat ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc
     7441 cctaactccg cccagttccg cccattctcc gccccatggc tgactaattt tttttattta
     7501 tgcagaggcc gaggccgcct ctgcctctga gctattccag aagtagtgag gaggcttttt
     7561 tggaggccta ggcttttgca aaaagctccc gggagcttgt atatccattt tcggatctga
     7621 tcagcacgtg ttgacaatta atcatcggca tagtatatcg gcatagtata atacgacaag
     7681 gtgaggaact aaaccatggc caagttgacc agtgccgttc cggtgctcac cgcgcgcgac
     7741 gtcgccggag cggtcgagtt ctggaccgac cggctcgggt tctcccggga cttcgtggag
     7801 gacgacttcg ccggtgtggt ccgggacgac gtgaccctgt tcatcagcgc ggtccaggac
     7861 caggtggtgc cggacaacac cctggcctgg gtgtgggtgc gcggcctgga cgagctgtac
     7921 gccgagtggt cggaggtcgt gtccacgaac ttccgggacg cctccgggcc ggccatgacc
     7981 gagatcggcg agcagccgtg ggggcgggag ttcgccctgc gcgacccggc cggcaactgc
     8041 gtgcacttcg tggccgagga gcaggactga cacgtgctac gagatttcga ttccaccgcc
     8101 gccttctatg aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc
     8161 cagcgcgggg atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat
     8221 aatggttaca aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg
     8281 cattctagtt gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg
     8341 acctctagct agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
     8401 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
     8461 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     8521 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     8581 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
     8641 cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac
     8701 gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg
     8761 ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca
     8821 agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc
     8881 tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc
     8941 ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag
     9001 gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc
     9061 ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca
     9121 gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg
     9181 aagtggtggc ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg
     9241 aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct
     9301 ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     9361 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     9421 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     9481 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     9541 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     9601 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     9661 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     9721 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     9781 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     9841 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     9901 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     9961 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
    10021 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
    10081 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
    10141 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
    10201 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
    10261 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
    10321 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
    10381 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
    10441 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
    10501 tttccccgaa aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt
    10561 gcactctcag tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt
    10621 gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct
    10681 tgaccgacaa ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg
    10741 tacgggccag atatacgcgt tgacattgat tattgactag ttattaatag taatcaatta
    10801 cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg
    10861 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc
    10921 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa
    10981 ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca
    11041 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta
    11101 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt
    11161 acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg
    11221 acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca
    11281 actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca
    11341 gcgcgttttg cctgtactgg gtctctctgg ttagaccaga tctgagcctg ggagctctct
    11401 ggctaactag ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta
    11461 gtgtgtgccc gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca
    11521 gtgtggaaaa tctctagcag tggcgcccga acagggactt gaaagcgaaa gggaaaccag
    11581 aggagctctc tcgacgcagg actcggcttg ctgaagcgcg cacggcaaga ggcgaggggc
    11641 ggcgactggt gagtacgcca aaaattttga ctagcggagg ctagaaggag agagatgggt
    11701 gcgagagcgt cagtattaag cgggggagaa ttagatcgcg atgggaaaaa attcggttaa
    11761 ggccaggggg aaagaaaaaa tataaattaa aacatatagt atgggcaagc agggagctag
    11821 aacgattcgc agttaatcct ggcctgttag aaacatcaga aggctgtaga caaatactgg
    11881 gacagctaca accatccctt cagacaggat cagaagaact tagatcatta tataatacag
    11941 tagcaaccct ctattgtgtg catcaaagga tagagataaa agacaccaag gaagctttag
    12001 acaagataga ggaagagcaa aacaaaagta agaccaccgc acagcaagcg gccgctgatc
    12061 ttcagacctg gaggaggaga tatgagggac aattggagaa gtgaattata taaatataaa
    12121 gtagtaaaaa ttgaaccatt aggagtagca cccaccaagg caaagagaag agtggtgcag
    12181 agagaaaaaa gagcagtggg aataggagct ttgttccttg ggttcttggg agcagcagga
    12241 agcactatgg gcgcagcgtc aatgacgctg acggtacagg ccagacaatt attgtctggt
    12301 atagtgcagc agcagaacaa tttgctgagg gctattgagg cgcaacagca tctgttgcaa
    12361 ctcacagtct ggggcatcaa gcagctccag gcaagaatcc tggctgtgga aagataccta
    12421 aaggatcaac agctcctggg gatttggggt tgctctggaa aactcatttg caccactgct
    12481 gtgccttgga atgctagttg gagtaataaa tctctggaac agatttggaa tcacacgacc
    12541 tggatggagt gggacagaga aattaacaat tacacaagct taatacactc cttaattgaa
    12601 gaatcgcaaa accagcaaga aaagaatgaa caagaattat tggaattaga taaatgggca
    12661 agtttgtgga attggtttaa cataacaaat tggctgtggt atataaaatt attcataatg
    12721 atagtaggag gcttggtagg tttaagaata gtttttgctg tactttctat agtgaataga
    12781 gttaggcagg gatattcacc attatcgttt cagacccacc tcccaacccc gaggggaccc
    12841 gacaggcccg aaggaataga agaagaaggt ggagagagag acagagacag atccattcga
    12901 ttagtgaacg gatcggcact gcgtgcgcca attctgcaga caaatggcag tattcatcca
    12961 caattttaaa agaaaagggg ggattggggg gtacagtgca ggggaaagaa tagtagacat
    13021 aatagcaaca gacatacaaa ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt
    13081 tcgggtttat tacagggaca gcagagatcc agtttggtta attaaggtac c