pDIRECT_26G

LOCUS       pDIRECT_26G               16777 bp ds-DNA     circular SYN 06-AUG-2018
DEFINITION  Direct Cloning, Type: T-DNA, Engineering Reagent: ZmUbi:TaCas9_dead 
            + TaU6:gRNA , Plant Selection: PvUbi2:bar.
ACCESSION   .
VERSION     .
KEYWORDS    pDIRECT_26G
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 16777)
  AUTHORS   Cermak T, Curtin SJ, Gil-Humanes J, Cegan R, Kono TJY, Konecna E, 
            Belanto JJ, Starker CG, Mathre JW, Greenstein RL, Voytas DF
  TITLE     A multi-purpose toolkit to enable advanced genome engineering in 
            plants.
  JOURNAL   Plant Cell. 2017 May 18. pii: tpc.00922.2016. doi: 
            10.1105/tpc.16.00922.
  PUBMED    28522548
REFERENCE   2  (bases 1 to 16777)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 6, 2018 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..16777
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     CDS             1233..1862
                     /codon_start=1
                     /product="stability protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 StaA
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
     CDS             2291..3364
                     /codon_start=1
                     /product="replication protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 RepA
                     /translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
                     QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
                     KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
                     GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
                     ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
                     LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
                     LVMRYRNLIEGEASAGS"
     rep_origin      3430..3624
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     primer_bind     complement(3860..3882)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    3968..4108
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     4023..4042
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(4123..4140)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4294..4882)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4374..4393)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4969..5763)
                     /codon_start=1
                     /gene="aphA-3"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
                     TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
                     EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
                     PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
                     FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
     misc_feature    6188..6212
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     polyA_signal    6290..6464
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     CDS             complement(6471..7022)
                     /codon_start=1
                     /gene="Streptomyces hygroscopicus bar"
                     /product="phosphinothricin acetyltransferase"
                     /label=BlpR
                     /note="confers resistance to bialophos or phosphinothricin"
                     /translation="MSPERRPADIRRATEADMPAVCTIVNHYIETSTVNFRTEPQEPQE
                     WTDDLVRLRERYPWLVAEVDGEVAGIAYAGPWKARNAYDWTAESTVYVSPRHQRTGLGS
                     TLYTHLLKSLEAQGFKSVVAVIGLPNDPSVRMHEALGYAPRGMLRAAGFKHGNWHDVGF
                     WQLDFSLPVPPRPVLPVTEI"
     protein_bind    8962..8983
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        8998..9028
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    9036..9052
                     /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)."
     primer_bind     9060..9076
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             11132..15235
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the H840A mutation in the HNH 
                     catalytic domain"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             15236..15283
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             complement(15920..16225)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVL
                     QRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSGTCL"
     primer_bind     16090..16106
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     16116..16135
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        16116..16134
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(16182..16202)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(16182..16200)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(16221..16237)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    16245..16261
                     /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(16269..16299)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    16314..16335
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     misc_RNA        16374..16449
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     complement(16466..16482)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    16685..16709
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
ORIGIN
        1 tccatttgta tgtgcatgcc aaccacaggg ttcccctcgg gatcaaagta ctttgatcca
       61 acccctccgc tgctatagtg cagtcggctt ctgacgttca gtgcagccgt cttctgaaaa
      121 cgacatgtcg cacaagtcct aagttacgcg acaggctgcc gccctgccct tttcctggcg
      181 ttttcttgtc gcgtgtttta gtcgcataaa gtagaatact tgcgactaga accggagaca
      241 ttacgccatg aacaagagcg ccgccgctgg cctgctgggc tatgcccgcg tcagcaccga
      301 cgaccaggac ttgaccaacc aacgggccga actgcacgcg gccggctgca ccaagctgtt
      361 ttccgagaag atcaccggca ccaggcgcga ccgcccggag ctggccagga tgcttgacca
      421 cctacgccct ggcgacgttg tgacagtgac caggctagac cgcctggccc gcagcacccg
      481 cgacctactg gacattgccg agcgcatcca ggaggccggc gcgggcctgc gtagcctggc
      541 agagccgtgg gccgacacca ccacgccggc cggccgcatg gtgttgaccg tgttcgccgg
      601 cattgccgag ttcgagcgtt ccctaatcat cgaccgcacc cggagcgggc gcgaggccgc
      661 caaggcccga ggcgtgaagt ttggcccccg ccctaccctc accccggcac agatcgcgca
      721 cgcccgcgag ctgatcgacc aggaaggccg caccgtgaaa gaggcggctg cactgcttgg
      781 cgtgcatcgc tcgaccctgt accgcgcact tgagcgcagc gaggaagtga cgcccaccga
      841 ggccaggcgg cgcggtgcct tccgtgagga cgcattgacc gaggccgacg ccctggcggc
      901 cgccgagaat gaacgccaag aggaacaagc atgaaaccgc accaggacgg ccaggacgaa
      961 ccgtttttca ttaccgaaga gatcgaggcg gagatgatcg cggccgggta cgtgttcgag
     1021 ccgcccgcgc acggctcaac cgtgcggctg catgaaatcc tggccggttt gtctgatgcc
     1081 aagctggcgg cctggccggc cagcttggcc gctgaagaaa ccgagcgccg ccgtctaaaa
     1141 aggtgatgtg tatttgagta aaacagcttg cgtcatgcgg tcgctgcgta tatgatgcga
     1201 tgagtaaata aacaaatacg caaggggaac gcatgaaggt tatcgctgta cttaaccaga
     1261 aaggcgggtc aggcaagacg accatcgcaa cccatctagc ccgcgccctg caactcgccg
     1321 gggccgatgt tctgttagtc gattccgatc cccagggcag tgcccgcgat tgggcggccg
     1381 tgcgggaaga tcaaccgcta accgttgtcg gcatcgaccg cccgacgatt gaccgcgacg
     1441 tgaaggccat cggccggcgc gacttcgtag tgatcgacgg agcgccccag gcggcggact
     1501 tggctgtgtc cgcgatcaag gcagccgact tcgtgctgat tccggtgcag ccaagccctt
     1561 acgacatatg ggccaccgcc gacctggtgg agctggttaa gcagcgcatt gaggtcacgg
     1621 atggaaggct acaagcggcc tttgtcgtgt cgcgggcgat caaaggcacg cgcatcggcg
     1681 gtgaggttgc cgaggcgctg gccgggtacg agctgcccat tcttgagtcc cgtatcacgc
     1741 agcgcgtgag ctacccaggc actgccgccg ccggcacaac cgttcttgaa tcagaacccg
     1801 agggcgacgc tgcccgcgag gtccaggcgc tggccgctga aattaaatca aaactcattt
     1861 gagttaatga ggtaaagaga aaatgagcaa aagcacaaac acgctaagtg ccggccgtcc
     1921 gagcgcacgc agcagcaagg ctgcaacgtt ggccagcctg gcagacacgc cagccatgaa
     1981 gcgggtcaac tttcagttgc cggcggagga tcacaccaag ctgaagatgt acgcggtacg
     2041 ccaaggcaag accattaccg agctgctatc tgaatacatc gcgcagctac cagagtaaat
     2101 gagcaaatga ataaatgagt agatgaattt tagcggctaa aggaggcggc atggaaaatc
     2161 aagaacaacc aggcaccgac gccgtggaat gccccatgtg tggaggaacg ggcggttggc
     2221 caggcgtaag cggctgggtt gcctgccggc cctgcaatgg cactggaacc cccaagcccg
     2281 aggaatcggc gtgagcggtc gcaaaccatc cggcccggta caaatcggcg cggcgctggg
     2341 tgatgacctg gtggagaagt tgaaggccgc gcaggccgcc cagcggcaac gcatcgaggc
     2401 agaagcacgc cccggtgaat cgtggcaagc ggccgctgat cgaatccgca aagaatcccg
     2461 gcaaccgccg gcagccggtg cgccgtcgat taggaagccg cccaagggcg acgagcaacc
     2521 agattttttc gttccgatgc tctatgacgt gggcacccgc gatagtcgca gcatcatgga
     2581 cgtggccgtt ttccgtctgt cgaagcgtga ccgacgagct ggcgaggtga tccgctacga
     2641 gcttccagac gggcacgtag aggtttccgc agggccggcc ggcatggcca gtgtgtggga
     2701 ttacgacctg gtactgatgg cggtttccca tctaaccgaa tccatgaacc gataccggga
     2761 agggaaggga gacaagcccg gccgcgtgtt ccgtccacac gttgcggacg tactcaagtt
     2821 ctgccggcga gccgatggcg gaaagcagaa agacgacctg gtagaaacct gcattcggtt
     2881 aaacaccacg cacgttgcca tgcagcgtac gaagaaggcc aagaacggcc gcctggtgac
     2941 ggtatccgag ggtgaagcct tgattagccg ctacaagatc gtaaagagcg aaaccgggcg
     3001 gccggagtac atcgagatcg agctagctga ttggatgtac cgcgagatca cagaaggcaa
     3061 gaacccggac gtgctgacgg ttcaccccga ttactttttg atcgatcccg gcatcggccg
     3121 ttttctctac cgcctggcac gccgcgccgc aggcaaggca gaagccagat ggttgttcaa
     3181 gacgatctac gaacgcagtg gcagcgccgg agagttcaag aagttctgtt tcaccgtgcg
     3241 caagctgatc gggtcaaatg acctgccgga gtacgatttg aaggaggagg cggggcaggc
     3301 tggcccgatc ctagtcatgc gctaccgcaa cctgatcgag ggcgaagcat ccgccggttc
     3361 ctaatgtacg gagcagatgc tagggcaaat tgccctagca ggggaaaaag gtcgaaaagg
     3421 cctctttcct gtggatagca cgtacattgg gaacccaaag ccgtacattg ggaaccggaa
     3481 cccgtacatt gggaacccaa agccgtacat tgggaaccgg tcacacatgt aagtgactga
     3541 tataaaagag aaaaaaggcg atttttccgc ctaaaactct ttaaaactta ttaaaactct
     3601 taaaacccgc ctggcctgtg cataactgtc tggccagcgc acagcccaag agctgcaaaa
     3661 agcgcctacc cttcggtcgc tgcgctccct acgccccgcc gcttcgcgtc ggcctatcgc
     3721 ggccgctggc cgctcaaaaa tggctggcct acggccaggc aatctaccag ggcgcggaca
     3781 agccgcgccg tcgccactcg accgccggcg cccacatcaa ggcaccctgc ctcgcgcgtt
     3841 tcggtgatga cggtgaaaac ctctgacaca tgcagctccc ggaaacggtc acagcttgtc
     3901 tgtaagcgga tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt
     3961 gtcggggcgc agccatgacc cagtcacgta gcgatagcgg agtgtatact ggcttaacta
     4021 tgcggcatca gagcagattg tactgagagt gcaccatatg cggtgtgaaa taccgcacag
     4081 atgcgtaagg agaaaatacc gcatcaggcc ctcttccgct tcctcgctca ctgactcgct
     4141 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
     4201 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
     4261 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     4321 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     4381 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     4441 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     4501 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     4561 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     4621 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     4681 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt
     4741 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     4801 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     4861 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     4921 gtggaacgaa aactcacgtt aagggatttt ggtcatgcat tctaggtact aaaacaattc
     4981 atccagtaaa atataatatt ttattttctc ccaatcaggc ttgatcccca gtaagtcaaa
     5041 aaatagctcg acatactgtt cttccccgat atcctccctg atcgaccgga cgcagaaggc
     5101 aatgtcatac cacttgtccg ccctgccgct tctcccaaga tcaataaagc cacttacttt
     5161 gccatctttc acaaagatgt tgctgtctcc caggtcgccg tgggaaaaga caagttcctc
     5221 ttcgggcttt tccgtcttta aaaaatcata cagctcgcgc ggatctttaa atggagtgtc
     5281 ttcttcccag ttttcgcaat ccacatcggc cagatcgtta ttcagtaagt aatccaattc
     5341 ggctaagcgg ctgtctaagc tattcgtata gggacaatcc gatatgtcga tggagtgaaa
     5401 gagcctgatg cactccgcat acagctcgat aatcttttca gggctttgtt catcttcata
     5461 ctcttccgag caaaggacgc catcggcctc actcatgagc agattgctcc agccatcatg
     5521 ccgttcaaag tgcaggacct ttggaacagg cagctttcct tccagccata gcatcatgtc
     5581 cttttcccgt tccacatcat aggtggtccc tttataccgg ctgtccgtca tttttaaata
     5641 taggttttca ttttctccca ccagcttata taccttagca ggagacattc cttccgtatc
     5701 ttttacgcag cggtattttt cgatcagttt tttcaattcc ggtgatattc tcattttagc
     5761 catttattat ttccttcctc ttttctacag tatttaaaga taccccaaga agctaattat
     5821 aacaagacga actccaattc actgttcctt gcattctaaa accttaaata ccagaaaaca
     5881 gctttttcaa agttgttttc aaagttggcg tataacatag tatcgacgga gccgattttg
     5941 aaaccgcggt gatcacaggc agcaacgctc tgtcatcgtt acaatcaaca tgctaccctc
     6001 cgcgagatca tccgtgtttc aaacccggca gcttagttgc cgttcttccg aatagcatcg
     6061 gtaacatgag caaagtctgc cgccttacaa cggctctccc gctgacgccg tcccggactg
     6121 atgggctgcc tgtatcgagt ggtgattttg tgccgagctg ccggtcgggg agctgttggc
     6181 tggctggtgg caggatatat tgtggtgtaa acaaattgac gcttagacaa cttaataaca
     6241 cattgcggac gtttttaatg tactgaatta acgccgaatt aattcggggg atctggattt
     6301 tagtactgga ttttggtttt aggaattaga aattttattg atagaagtat tttacaaata
     6361 caaatacata ctaagggttt cttatatgct caacacatga gcgaaaccct ataggaaccc
     6421 taattccctt atctgggaac tactcacaca ttattatgga gaaactcgag tcaaatctcg
     6481 gtgacgggca ggaccggacg gggcggtacc ggcaggctga agtccagctg ccagaaaccc
     6541 acgtcatgcc agttcccgtg cttgaagccg gccgcccgca gcatgccgcg gggggcatat
     6601 ccgagcgcct cgtgcatgcg cacgctcggg tcgttgggca gcccgatgac agcgaccacg
     6661 ctcttgaagc cctgtgcctc cagggacttc agcagatggg tgtagagcgt ggagcccagt
     6721 cccgtccgct ggtggcgggg gctgacgtac acggtggact cggccgtcca gtcgtaggcg
     6781 ttgcgtgcct tccagggacc cgcgtaggcg atgccggcga cctcgccgtc cacctcggcg
     6841 acgagccagg gatagcgctc ccgcagacgg acgaggtcgt ccgtccactc ctgcggttcc
     6901 tgcggctcgg tacggaagtt gaccgtgctt gtctcgatgt agtggttgac gatggtgcag
     6961 accgccggca tgtccgcctc ggtggcacgg cggatgtcgg ccgggcgtcg ttctgggctc
     7021 ataagcttct gcaaaagaga accagacaac agggtaagtg cctagcagta aacaaacaga
     7081 actcatcaca agcaaacagc aacatcatat tcataccaac aggtcatgtg tgttcatcac
     7141 atcattagta ctaagcatgc catcatccaa gtatatcaaa gtaagggcaa agagcatgca
     7201 tgatcatcag gtgcacaaaa gaatcatcaa attgtagcag tacatatctt catctatcat
     7261 gcatatctat ccataacagg acgatgcatg ttgaccaggt aaaagctaca ggatcctata
     7321 ggaacagcag cgtatatatc ttcaccaatc ttagcatctt aatcatgtgg cacatgcagt
     7381 ttcaatttaa gcacatgagc tagttgatta tgaggtacca gagaatcatc aaactgatgt
     7441 agcagcatat atcctcatct atcatgcaat ctaatctaat ctaactaaac aggaaaggtg
     7501 tgctattcag ttaaaagcta ccgcatcata tacaaacggc agcataagaa aaagcataat
     7561 catcttaatc atgcaacaaa cgcagattca taataagccc aagagctagc ttgtgatgat
     7621 cttattctac tctgatctac agcaatcaga taacgaccta accttgcaca tggcaacaaa
     7681 acaatcgatc ggacgaatca gttgtttgtt cctagctagc accatcgaac cagataatag
     7741 atgcacgtac agatcccgaa aacgaaccca aaaacagggc agacctagct gaacctaggc
     7801 agcgacccag cagatcgtga gaacgatctc atctacgaac agcctagaag caaccccacg
     7861 attcccggac aaacgaccta aaatccccac aaatcacatg agcatgacag gataaacagc
     7921 ggaaccgatc agatctacac gaaaacccca cctcccagcc acccacgatc aggaaacacg
     7981 cggatctagc atgatttcgt caacgcctca gcctagttcc tagccacaga ccaagcagaa
     8041 ccaccaaacc acgccgagcg aggagatggg gcaagaggac gggggagacg atcgccgtac
     8101 cttgaagcgg gggaaggatc gccgagggtc gcgaggagag caatttggat ttggaacccg
     8161 ggggttgtgc gctccgaacg atgagacgat gtgagattgt gggaagaggc gcggagggcc
     8221 ctgtatttat gggctgcgac ggggggagga gaggtgggga gggttgggga aggaatcccc
     8281 cacccgtgcc gtgacggttc cgggccgtgt gagaggagcc cgctcgtctc cgccacgcaa
     8341 tttccgcgat cggagcggag ctttcgagag gcggctggat ggttggtggc cgttagattt
     8401 gtagacgccg ttaacgcctc gcctccaccg ggaagagttt tgagcagccg cttatgacaa
     8461 tggcttaacg acgttagacg gagcgttagt ggcaggccat gatagaatag acgtattgca
     8521 atgggattat aattaaataa ataagaatat aataagatat ggcaagtcgg cactcatgac
     8581 atggtcttcg aaatgatagt gctcactttc ttagccgaga aagttgacgc gactgattta
     8641 gaagttaaga tattatttct ctcttctttt ctttcctcgt catataagga tgaaataaac
     8701 tttagagatt gccggtagtg attttggatt tcggcgatca ggcttggttt gccggtttcg
     8761 gacggtgtgc cttaggccac ccgcagtgta tcttgtaatg ttcaaccgat aagcaagggt
     8821 ggggctcaag caagtagtaa acaactatgt caaatgtcac catggttatg gtcttgttta
     8881 gttggcttct tggccgattc attaatgcag ctggcacgac aggtttcccg actggaaagc
     8941 gggcagtgag cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt
     9001 acactttatg cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac
     9061 aggaaacagc tatgacatga ttacgaattc gagctcggta cccggggatc tgcaggcgcg
     9121 cctgcagtgc agcgtgaccc ggtcgtgccc ctctctagag ataatgagca ttgcatgtct
     9181 aagttataaa aaattaccac atattttttt tgtcacactt gtttgaagtg cagtttatct
     9241 atctttatac atatatttaa actttactct acgaataata taatctatag tactacaata
     9301 atatcagtgt tttagagaat catataaatg aacagttaga catggtctaa aggacaattg
     9361 agtattttga caacaggact ctacagtttt atctttttag tgtgcatgtg ttctcctttt
     9421 tttttgcaaa tagcttcacc tatataatac ttcatccatt ttattagtac atccatttag
     9481 ggtttagggt taatggtttt tatagactaa tttttttagt acatctattt tattctattt
     9541 tagcctctaa attaagaaaa ctaaaactct attttagttt ttttatttaa taatttagat
     9601 ataaaataga ataaaataaa gtgactaaaa attaaacaaa taccctttaa gaaattaaaa
     9661 aaactaagga aacatttttc ttgtttcgag tagataatgc cagcctgtta aacgccgtcg
     9721 acgagtctaa cggacaccaa ccagcgaacc agcagcgtcg cgtcgggcca agcgaagcag
     9781 acggcacggc atctctgtcg ctgcctctgg acccctctcg agagttccgc tccaccgttg
     9841 gacttgctcc gctgtcggca tccagaaatt gcgtggcgga gcggcagacg tgagccggca
     9901 cggcaggcgg cctcctcctc ctctcacggc accggcagct acgggggatt cctttcccac
     9961 cgctccttcg ctttcccttc ctcgcccgcc gtaataaata gacaccccct ccacaccctc
    10021 tttccccaac ctcgtgttgt tcggagcgca cacacacaca accagatctc ccccaaatcc
    10081 acccgtcggc acctccgctt caaggtacgc cgctcgtcct cccccccccc ctctctacct
    10141 tctctagatc ggcgttccgg tccatggtta gggcccggta gttctacttc tgttcatgtt
    10201 tgtgttagat ccgtgtttgt gttagatccg tgctgctagc gttcgtacac ggatgcgacc
    10261 tgtacgtcag acacgttctg attgctaact tgccagtgtt tctctttggg gaatcctggg
    10321 atggctctag ccgttccgca gacgggatcg atttcatgat tttttttgtt tcgttgcata
    10381 gggtttggtt tgcccttttc ctttatttca atatatgccg tgcacttgtt tgtcgggtca
    10441 tcttttcatg cttttttttg tcttggttgt gatgatgtgg tctggttggg cggtcgttct
    10501 agatcggagt agaattaatt ctgtttcaaa ctacctggtg gatttattaa ttttggatct
    10561 gtatgtgtgt gccatacata ttcatagtta cgaattgaag atgatggatg gaaatatcga
    10621 tctaggatag gtatacatgt tgatgcgggt tttactgatg catatacaga gatgcttttt
    10681 gttcgcttgg ttgtgatgat gtggtgtggt tgggcggtcg ttcattcgtt ctagatcgga
    10741 gtagaatact gtttcaaact acctggtgta tttattaatt ttggaactgt atgtgtgtgt
    10801 catacatctt catagttacg agtttaagat ggatggaaat atcgatctag gataggtata
    10861 catgttgatg tgggttttac tgatgcatat acatgatggc atatgcagca tctattcata
    10921 tgctctaacc ttgagtacct atctattata ataaacaagt atgttttata attattttga
    10981 tcttgatata cttggatgat ggcatatgca gcagctatat gtggattttt ttagccctgc
    11041 cttcatacgc tatttatttg cttggtactg tttcttttgt cgatgctcac cctgttgttt
    11101 ggtgttactt cctgcagggc gatctattcg aatggacaag aagtactcga tcggcctcgc
    11161 catcgggacg aactcagttg gctgggccgt gatcaccgac gagtacaagg tgccctctaa
    11221 gaagttcaag gtcctgggga acaccgaccg ccattccatc aagaagaacc tcatcggcgc
    11281 tctcctgttc gacagcgggg agaccgctga ggctacgagg ctcaagagaa ccgctaggcg
    11341 ccggtacacg agaaggaaga acaggatctg ctacctccaa gagattttct ccaacgagat
    11401 ggccaaggtt gacgattcat tcttccaccg cctggaggag tctttcctcg tggaggagga
    11461 taagaagcac gagcggcatc ccatcttcgg caacatcgtg gacgaggttg cctaccacga
    11521 gaagtaccct acgatctacc atctgcggaa gaagctcgtg gactccaccg ataaggcgga
    11581 cctcagactg atctacctcg ctctggccca catgatcaag ttccgcggcc atttcctgat
    11641 cgagggggat ctcaacccag acaacagcga tgttgacaag ctgttcatcc aactcgtgca
    11701 gacctacaac caactcttcg aggagaaccc gatcaacgcc tctggcgtgg acgcgaaggc
    11761 tatcctgtcc gcgaggctct cgaagtccag gaggctggag aacctgatcg ctcagctccc
    11821 aggcgagaag aagaacggcc tgttcgggaa cctcatcgct ctcagcctgg ggctcacccc
    11881 gaacttcaag tcgaacttcg atctcgctga ggacgccaag ctgcaactct ccaaggacac
    11941 ctacgacgat gacctcgata acctcctggc ccagatcggc gatcaatacg cggacctgtt
    12001 cctcgctgcc aagaacctgt cggacgccat cctcctgtca gatatcctcc gcgtgaacac
    12061 cgagatcacg aaggctccac tctctgcctc catgatcaag cgctacgacg agcaccatca
    12121 ggatctgacc ctcctgaagg cgctggtccg ccaacagctc ccggagaagt acaaggagat
    12181 tttcttcgat cagtcgaaga acggctacgc tgggtacatc gacggcgggg cctcacaaga
    12241 ggagttctac aagttcatca agccaatcct ggagaagatg gacggcacgg aggagctcct
    12301 ggtgaagctc aacagggagg acctcctgcg gaagcagaga accttcgata acggcagcat
    12361 cccccaccaa atccatctcg gggagctgca cgccatcctg agaaggcaag aggacttcta
    12421 ccctttcctc aaggataacc gggagaagat cgagaagatc ctgaccttca gaatcccata
    12481 ctacgtcggc cctctcgcgc gggggaactc aagattcgct tggatgaccc gcaagtctga
    12541 ggagaccatc acgccgtgga acttcgagga ggtggtggac aagggcgcta gcgctcagtc
    12601 gttcatcgag aggatgacca acttcgacaa gaacctgccc aacgagaagg tgctccctaa
    12661 gcactcgctc ctgtacgagt acttcaccgt ctacaacgag ctcacgaagg tgaagtacgt
    12721 caccgagggc atgcgcaagc cagcgttcct gtccggggag cagaagaagg ctatcgtgga
    12781 cctcctgttc aagaccaacc ggaaggtcac ggttaagcaa ctcaaggagg actacttcaa
    12841 gaagatcgag tgcttcgatt cggtcgagat cagcggcgtt gaggaccgct tcaacgccag
    12901 cctcgggacc taccacgatc tcctgaagat catcaaggat aaggacttcc tggacaacga
    12961 ggagaacgag gatatcctgg aggacatcgt gctgaccctc acgctgttcg aggacaggga
    13021 gatgatcgag gagcgcctga agacgtacgc ccatctcttc gatgacaagg tcatgaagca
    13081 actcaagcgc cggagataca ccggctgggg gaggctgtcc cgcaagctca tcaacggcat
    13141 ccgggacaag cagtccggga agaccatcct cgacttcctg aagagcgatg gcttcgccaa
    13201 caggaacttc atgcaactga tccacgatga cagcctcacc ttcaaggagg atatccaaaa
    13261 ggctcaagtg agcggccagg gggactcgct gcacgagcat atcgcgaacc tcgctggctc
    13321 ccccgcgatc aagaagggca tcctccagac cgtgaaggtt gtggacgagc tcgtgaaggt
    13381 catgggccgg cacaagcctg agaacatcgt catcgagatg gccagagaga accaaaccac
    13441 gcagaagggg caaaagaact ctagggagcg catgaagcgc atcgaggagg gcatcaagga
    13501 gctggggtcc caaatcctca aggagcaccc agtggagaac acccaactgc agaacgagaa
    13561 gctctacctg tactacctcc agaacggcag ggatatgtac gtggaccaag agctggatat
    13621 caaccgcctc agcgattacg acgtcgatgc tatcgttccc cagtctttcc tgaaggatga
    13681 ctccatcgac aacaaggtcc tcaccaggtc ggacaagaac cgcggcaagt cagataacgt
    13741 tccatctgag gaggtcgtta agaagatgaa gaactactgg aggcagctcc tgaacgccaa
    13801 gctgatcacg caaaggaagt tcgacaacct caccaaggct gagagaggcg ggctctcaga
    13861 gctggacaag gccggcttca tcaagcggca gctggtcgag accagacaaa tcacgaagca
    13921 cgttgcgcaa atcctcgact ctcggatgaa cacgaagtac gatgagaacg acaagctgat
    13981 cagggaggtt aaggtgatca ccctgaagtc taagctcgtc tccgacttca ggaaggattt
    14041 ccagttctac aaggttcgcg agatcaacaa ctaccaccat gcccatgacg cttacctcaa
    14101 cgctgtggtc ggcaccgctc tgatcaagaa gtacccaaag ctggagtccg agttcgtgta
    14161 cggggactac aaggtttacg atgtgcgcaa gatgatcgcc aagtcggagc aagagatcgg
    14221 caaggctacc gccaagtact tcttctactc aaacatcatg aacttcttca agaccgagat
    14281 cacgctggcc aacggcgaga tccggaagag accgctcatc gagaccaacg gcgagacggg
    14341 ggagatcgtg tgggacaagg gcagggattt cgcgaccgtc cgcaaggttc tctccatgcc
    14401 ccaggtgaac atcgtcaaga agaccgaggt ccaaacgggc gggttctcaa aggagtctat
    14461 cctgcctaag cggaacagcg acaagctcat cgccagaaag aaggactggg acccaaagaa
    14521 gtacggcggg ttcgacagcc ctaccgtggc ctactcggtc ctggttgtgg cgaaggttga
    14581 gaagggcaag tccaagaagc tcaagagcgt gaaggagctc ctggggatca ccatcatgga
    14641 gaggtccagc ttcgagaaga acccaatcga cttcctggag gccaagggct acaaggaggt
    14701 gaagaaggac ctgatcatca agctcccgaa gtactctctc ttcgagctgg agaacggcag
    14761 gaagagaatg ctggcttccg ctggcgagct ccagaagggg aacgagctcg cgctgccaag
    14821 caagtacgtg aacttcctct acctggcttc ccactacgag aagctcaagg gcagcccgga
    14881 ggacaacgag caaaagcagc tgttcgtcga gcagcacaag cattacctcg acgagatcat
    14941 cgagcaaatc tccgagttca gcaagcgcgt gatcctcgcc gacgcgaacc tggataaggt
    15001 cctctccgcc tacaacaagc accgggacaa gcccatcaga gagcaagcgg agaacatcat
    15061 ccatctcttc accctgacga acctcggcgc tcctgctgct ttcaagtact tcgacaccac
    15121 gatcgatcgg aagagataca cctccacgaa ggaggtcctg gacgcgaccc tcatccacca
    15181 gtcgatcacc ggcctgtacg agacgaggat cgacctctca caactcggcg gggataagag
    15241 acccgcagca accaagaagg cagggcaagc aaagaagaag aagtgactcg agatatgaag
    15301 atgaagatga aatatttggt gtgtcaaata aaaagcttgt gtgcttaagt ttgtgttttt
    15361 ttcttggctt gttgtgttat gaatttgtgg ctttttctaa tattaaatga atgtaagatc
    15421 acattataat gaataaacaa atgtttctat aatccattgt gaatgttttg ttggatctct
    15481 tctgcagcat ataactactg tatgtgctat ggtatggact atggaatatg attaaagata
    15541 aggagctccg gtgacggacg accaagcccg ttattctgac agttctggtg ctcaacacat
    15601 ttatatttat caaggagcac attgttactc actgctagga gggaatcgaa ctaggaatat
    15661 tgatcagagg aactacgaga gagctgaaga taactgccct ctagctctca ctgatctggg
    15721 tcgcatagtg agatgcagcc cacgtgagtt cagcaacggt ctagcgctgg gcttttaggc
    15781 ccgcatgatc gggcttttgt cgggtggtcg acgtgttcac gattggggag agcaacgcag
    15841 cagttcctct tagtttagtc ccacctcgcc tgtccagcag agttctgacc ggtttataaa
    15901 ctcgcttgct gcatcagact tagaggcagg tgccgctaca gggcgcgtcc cattcgccat
    15961 tcaggctgcg caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc
    16021 tggcgaaagg gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt
    16081 cacgacgttg taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat
    16141 tgggtaccgg gccccccctc gaggtcctcc agcttttgtt ccctttagtg agggttaatt
    16201 gcgcgctcgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta tccgctcaca
    16261 attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc ctaatgagtg
    16321 agctaactca cattaattgc gttgcgctca ctgcccgctt tccacctgcg gtcgttttag
    16381 agctagaaat agcaagttaa aataaggcta gtccgttatc aacttgaaaa agtggcaccg
    16441 agtcggtgct ttttttcccg gtggcactgg ccgtcgtttt acaacgtcgt gactgggaaa
    16501 accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc agctggcgta
    16561 atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg aatggcgaat
    16621 gctagagcag cttgagcttg gatcagattg tcgtttcccg ccttcagttt aaactatcag
    16681 tgtttgacag gatatattgg cgggtaaacc taagagaaaa gagcgtttat tagaataatc
    16741 ggatatttaa aagggcgtga aaaggtttat ccgttcg
//