pAAVS1-NC-CRISPRi (Gen3)

LOCUS       pAAVS1-NC-CRISPRi (Gen3)             11437 bp ds-DNA     circular SYN 06-AUG-2018
DEFINITION  Constitutive CRISPR interference (CRISPRi) knock in construct into 
            the AAVS1 locus.
ACCESSION   .
VERSION     .
KEYWORDS    pAAVS1-NC-CRISPRi (Gen3)
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11437)
  AUTHORS   Mandegar MA, Huebsch N, Frolov EB, Shin E, Truong A, Olvera MP, Chan
            AH, Miyaoka Y, Holmes K, Spencer CI, Judge LM, Gordon DE, Eskildsen 
            TV, Villalta JE, Horlbeck MA, Gilbert LA, Krogan NJ, Sheikh SP, 
            Weissman JS, Qi LS, So PL, Conklin BR
  TITLE     CRISPR Interference Efficiently Induces Specific and Reversible Gene
            Silencing in Human iPSCs.
  JOURNAL   Cell Stem Cell. 2016 Apr 7;18(4):541-53. doi: 
            10.1016/j.stem.2016.01.022. Epub 2016 Mar 10.
  PUBMED    26971820
REFERENCE   2  (bases 1 to 11437)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 6, 2018 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..11437
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     misc_feature    1..804
                     /label=HA-L
                     /note="left homology arm from the adeno-associated virus 
                     integration site (AAVS1) within intron 1 of the human 
                     PPP1R12C gene"
     protein_bind    806..839
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    846..871
                     /label=SA
                     /note="splice acceptor site"
     CDS             895..948
                     /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"
     regulatory      964..973
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             970..1773
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGDDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1033..1052)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1643..1662
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(1793..1810)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1799..2023
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     enhancer        2029..2408
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2410..2687
                     /label=chicken beta-actin promoter
     intron          2689..3706
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     3630..3652
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     3714..3733
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      3765..3774
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             3801..3986
                     /codon_start=1
                     /gene="ZNF10"
                     /gene_synonym="KOX1"
                     /product="Kruppel-associated box (KRAB) transcriptional 
                     repression domain from the human zinc finger protein ZNF10 
                     (Margolin et al., 1994)"
                     /label=KRAB
                     /translation="RTLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGY
                     QLTKPDVILRLEKGEEP"
     CDS             4008..8111
                     /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             8115..8141
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             8160..8180
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8187..8207
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(8338..8357)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_site      8403..8458
                     /label=-beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal"
     primer_bind     complement(8457..8476)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     protein_bind    8807..8840
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    8881..9717
                     /label=HA-R
                     /note="right homology arm from the adeno-associated virus 
                     integration site (AAVS1) within intron 1 of the human 
                     PPP1R12C gene"
     rep_origin      complement(9765..10353)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(9845..9864)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(10524..11384)
                     /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     11147..11166
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 tgctttctct gaccagcatt ctctcccctg ggcctgtgcc gctttctgtc tgcagcttgt
       61 ggcctgggtc acctctacgg ctggcccaga tccttccctg ccgcctcctt caggttccgt
      121 cttcctccac tccctcttcc ccttgctctc tgctgtgttg ctgcccaagg atgctctttc
      181 cggagcactt ccttctcggc gctgcaccac gtgatgtcct ctgagcggat cctccccgtg
      241 tctgggtcct ctccgggcat ctctcctccc tcacccaacc ccatgccgtc ttcactcgct
      301 gggttccctt ttccttctcc ttctggggcc tgtgccatct ctcgtttctt aggatggcct
      361 tctccgacgg atgtctccct tgcgtcccgc ctccccttct tgtaggcctg catcatcacc
      421 gtttttctgg acaaccccaa agtaccccgt ctccctggct ttagccacct ctccatcctc
      481 ttgctttctt tgcctggaca ccccgttctc ctgtggattc gggtcacctc tcactccttt
      541 catttgggca gctcccctac cccccttacc tctctagtct gtgctagctc ttccagcccc
      601 ctgtcatggc atcttccagg ggtccgagag ctcagctagt cttcttcctc caacccgggc
      661 ccctatgtcc acttcaggac agcatgtttg ctgcctccag ggatcctgtg tccccgagct
      721 gggaccacct tatattccca gggccggtta atgtggctct ggttctgggt acttttatct
      781 gtcccctcca ccccacagtg gggcaataac ttcgtatagc atacattata cgaagttata
      841 agcttctgac ctcttctctt cctcccacag ggcctcgaga gatctggcag cggagagggc
      901 agaggaagtc ttctaacatg cggtgacgtg gaggagaatc ccggccctag gctcgagggt
      961 accgccacca tgggatcggc cattgaacaa gatggattgc acgcaggttc tccggccgct
     1021 tgggtggaga ggctattcgg ctatgactgg gcacaacaga caatcggctg ctctgatgcc
     1081 gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt ttgtcaagac cgacctgtcc
     1141 ggtgccctga atgaactgca ggacgaggca gcgcggctat cgtggctggc cacgacgggc
     1201 gttccttgcg cagctgtgct cgacgttgtc actgaagcgg gaagggactg gctgctattg
     1261 ggcgaagtgc cggggcagga tctcctgtca tctcaccttg ctcctgccga gaaagtatcc
     1321 atcatggctg atgcaatgcg gcggctgcat acgcttgatc cggctacctg cccattcgac
     1381 caccaagcga aacatcgcat cgagcgagca cgtactcgga tggaagccgg tcttgtcgat
     1441 caggatgatc tggacgaaga gcatcagggg ctcgcgccag ccgaactgtt cgccaggctc
     1501 aaggcgcgca tgcccgacgg cgatgatctc gtcgtgaccc atggcgatgc ctgcttgccg
     1561 aatatcatgg tggaaaatgg ccgcttttct ggattcatcg actgtggccg gctgggtgtg
     1621 gcggaccgct atcaggacat agcgttggct acccgtgata ttgctgaaga gcttggcggc
     1681 gaatgggctg accgcttcct cgtgctttac ggtatcgccg ctcccgattc gcagcgcatc
     1741 gccttctatc gccttcttga cgagttcttc tgagtttaaa cccgctgatc agcctcgact
     1801 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg
     1861 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg
     1921 agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg
     1981 gaagacaata gcaggcatgc tggggatgcg gtgggctcta tgggtctcga cattgattat
     2041 tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt
     2101 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
     2161 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
     2221 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
     2281 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc
     2341 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
     2401 ttaccatggt cgaggtgagc cccacgttct gcttcactct ccccatctcc cccccctccc
     2461 cacccccaat tttgtattta tttatttttt aattattttg tgcagcgatg ggggcggggg
     2521 gggggggggg gcgcgcgcca ggcggggcgg ggcggggcga ggggcggggc ggggcgaggc
     2581 ggagaggtgc ggcggcagcc aatcagagcg gcgcgctccg aaagtttcct tttatggcga
     2641 ggcggcggcg gcggcggccc tataaaaagc gaagcgcgcg gcgggcgggg agtcgctgcg
     2701 acgctgcctt cgccccgtgc cccgctccgc cgccgcctcg cgccgcccgc cccggctctg
     2761 actgaccgcg ttactcccac aggtgagcgg gcgggacggc ccttctcctc cgggctgtaa
     2821 ttagcgcttg gtttaatgac ggcttgtttc ttttctgtgg ctgcgtgaaa gccttgaggg
     2881 gctccgggag ggccctttgt gcggggggag cggctcgggg ggtgcgtgcg tgtgtgtgtg
     2941 cgtggggagc gccgcgtgcg gctccgcgct gcccggcggc tgtgagcgct gcgggcgcgg
     3001 cgcggggctt tgtgcgctcc gcagtgtgcg cgaggggagc gcggccgggg gcggtgcccc
     3061 gcggtgcggg gggggctgcg aggggaacaa aggctgcgtg cggggtgtgt gcgtgggggg
     3121 gtgagcaggg ggtgtgggcg cgtcggtcgg gctgcaaccc cccctgcacc cccctccccg
     3181 agttgctgag cacggcccgg cttcgggtgc ggggctccgt acggggcgtg gcgcggggct
     3241 cgccgtgccg ggcggggggt ggcggcaggt gggggtgccg ggcggggcgg ggccgcctcg
     3301 ggccggggag ggctcggggg aggggcgcgg cggcccccgg agcgccggcg gctgtcgagg
     3361 cgcggcgagc cgcagccatt gccttttatg gtaatcgtgc gagagggcgc agggacttcc
     3421 tttgtcccaa atctgtgcgg agccgaaatc tgggaggcgc cgccgcaccc cctctagcgg
     3481 gcgcggggcg aagcggtgcg gcgccggcag gaaggaaatg ggcggggagg gccttcgtgc
     3541 gtcgccgcgc cgccgtcccc ttctccctct ccagcctcgg ggctgtccgc ggggggacgg
     3601 ctgccttcgg gggggacggg gcagggcggg gttcggcttc tggcgtgtga ccggcggctc
     3661 tagagcctct gctaaccatg ttcatgcctt cttctttttc ctacagctcc tgggcaacgt
     3721 gctggttatt gtgctgtctc atcattttgg caaagaatgt cgacgccacc atggatgcta
     3781 agtcactaac tgcctggtcc cggacactgg tgaccttcaa ggatgtattt gtggacttca
     3841 ccagggagga gtggaagctg ctggacactg ctcagcagat cgtgtacaga aatgtgatgc
     3901 tggagaacta taagaacctg gtttccttgg gttatcagct tactaagcca gatgtgatcc
     3961 tccggttgga gaagggagaa gagcccggcg gttccggcgg agggtcgatg gacaagaagt
     4021 acagcatcgg cctggccatc ggcaccaact ctgtgggctg ggccgtgatc accgacgagt
     4081 acaaggtgcc cagcaagaaa ttcaaggtgc tgggcaacac cgaccggcac agcatcaaga
     4141 agaacctgat cggcgccctg ctgttcgaca gcggagaaac agccgaggcc acccggctga
     4201 agagaaccgc cagaagaaga tacaccagac ggaagaaccg gatctgctat ctgcaagaga
     4261 tcttcagcaa cgagatggcc aaggtggacg acagcttctt ccacagactg gaagagtcct
     4321 tcctggtgga agaggataag aagcacgagc ggcaccccat cttcggcaac atcgtggacg
     4381 aggtggccta ccacgagaag taccccacca tctaccacct gagaaagaaa ctggtggaca
     4441 gcaccgacaa ggccgacctg cggctgatct atctggccct ggcccacatg atcaagttcc
     4501 ggggccactt cctgatcgag ggcgacctga accccgacaa cagcgacgtg gacaagctgt
     4561 tcatccagct ggtgcagacc tacaaccagc tgttcgagga aaaccccatc aacgccagcg
     4621 gcgtggacgc caaggccatc ctgtctgcca gactgagcaa gagcagacgg ctggaaaatc
     4681 tgatcgccca gctgcccggc gagaagaaga atggcctgtt cggcaacctg attgccctga
     4741 gcctgggcct gacccccaac ttcaagagca acttcgacct ggccgaggat gccaaactgc
     4801 agctgagcaa ggacacctac gacgacgacc tggacaacct gctggcccag atcggcgacc
     4861 agtacgccga cctgtttctg gccgccaaga acctgtccga cgccatcctg ctgagcgaca
     4921 tcctgagagt gaacaccgag atcaccaagg cccccctgag cgcctctatg atcaagagat
     4981 acgacgagca ccaccaggac ctgaccctgc tgaaagctct cgtgcggcag cagctgcctg
     5041 agaagtacaa agagattttc ttcgaccaga gcaagaacgg ctacgccggc tacatcgatg
     5101 gcggagccag ccaggaagag ttctacaagt tcatcaagcc catcctggaa aagatggacg
     5161 gcaccgagga actgctcgtg aagctgaaca gagaggacct gctgcggaag cagcggacct
     5221 tcgacaacgg cagcatcccc caccagatcc acctgggaga gctgcacgcc attctgcggc
     5281 ggcaggaaga tttttaccca ttcctgaagg acaaccggga aaagatcgag aagatcctga
     5341 ccttccgcat cccctactac gtgggccctc tggccagggg aaacagcaga ttcgcctgga
     5401 tgaccagaaa gagcgaggaa accatcaccc cctggaactt cgaggaagtg gtggacaagg
     5461 gcgccagcgc ccagagcttc atcgagcgga tgaccaactt cgataagaac ctgcccaacg
     5521 agaaggtgct gcccaagcac agcctgctgt acgagtactt caccgtgtac aacgagctga
     5581 ccaaagtgaa atacgtgacc gagggaatga gaaagcccgc cttcctgagc ggcgagcaga
     5641 aaaaagccat cgtggacctg ctgttcaaga ccaaccggaa agtgaccgtg aagcagctga
     5701 aagaggacta cttcaagaaa atcgagtgct tcgactccgt ggaaatctcc ggcgtggaag
     5761 atcggttcaa cgcctccctg ggcacatacc acgatctgct gaaaattatc aaggacaagg
     5821 acttcctgga caatgaggaa aacgaggaca ttctggaaga tatcgtgctg accctgacac
     5881 tgtttgagga cagagagatg atcgaggaac ggctgaaaac ctatgcccac ctgttcgacg
     5941 acaaagtgat gaagcagctg aagcggcgga gatacaccgg ctggggcagg ctgagccgga
     6001 agctgatcaa cggcatccgg gacaagcagt ccggcaagac aatcctggat ttcctgaagt
     6061 ccgacggctt cgccaacaga aacttcatgc agctgatcca cgacgacagc ctgaccttta
     6121 aagaggacat ccagaaagcc caggtgtccg gccagggcga tagcctgcac gagcacattg
     6181 ccaatctggc cggcagcccc gccattaaga agggcatcct gcagacagtg aaggtggtgg
     6241 acgagctcgt gaaagtgatg ggccggcaca agcccgagaa catcgtgatc gaaatggcca
     6301 gagagaacca gaccacccag aagggacaga agaacagccg cgagagaatg aagcggatcg
     6361 aagagggcat caaagagctg ggcagccaga tcctgaaaga acaccccgtg gaaaacaccc
     6421 agctgcagaa cgagaagctg tacctgtact acctgcagaa tgggcgggat atgtacgtgg
     6481 accaggaact ggacatcaac cggctgtccg actacgatgt ggacgctatc gtgcctcaga
     6541 gctttctgaa ggacgactcc atcgataaca aagtgctgac tcggagcgac aagaaccggg
     6601 gcaagagcga caacgtgccc tccgaagagg tcgtgaagaa gatgaagaac tactggcgcc
     6661 agctgctgaa tgccaagctg attacccaga ggaagttcga caatctgacc aaggccgaga
     6721 gaggcggcct gagcgaactg gataaggccg gcttcatcaa gagacagctg gtggaaaccc
     6781 ggcagatcac aaagcacgtg gcacagatcc tggactcccg gatgaacact aagtacgacg
     6841 agaacgacaa actgatccgg gaagtgaaag tgatcaccct gaagtccaag ctggtgtccg
     6901 atttccggaa ggatttccag ttttacaaag tgcgcgagat caacaactac caccacgccc
     6961 acgacgccta cctgaacgcc gtcgtgggaa ccgccctgat caaaaagtac cctaagctgg
     7021 aaagcgagtt cgtgtacggc gactacaagg tgtacgacgt gcggaagatg atcgccaaga
     7081 gcgagcagga aatcggcaag gctaccgcca agtacttctt ctacagcaac atcatgaact
     7141 ttttcaagac cgagattacc ctggccaacg gcgagatccg gaagcggcct ctgatcgaga
     7201 caaacggcga aacaggcgag atcgtgtggg ataagggccg ggactttgcc accgtgcgga
     7261 aagtgctgtc tatgccccaa gtgaatatcg tgaaaaagac cgaggtgcag acaggcggct
     7321 tcagcaaaga gtctatcctg cccaagagga acagcgacaa gctgatcgcc agaaagaagg
     7381 actgggaccc taagaagtac ggcggcttcg acagccccac cgtggcctat tctgtgctgg
     7441 tggtggccaa agtggaaaag ggcaagtcca agaaactgaa gagtgtgaaa gagctgctgg
     7501 ggatcaccat catggaaaga agcagcttcg agaagaatcc catcgacttt ctggaagcca
     7561 agggctacaa agaagtgaaa aaggacctga tcatcaagct gcctaagtac tccctgttcg
     7621 agctggaaaa cggccggaag agaatgctgg cctctgccgg cgaactgcag aagggaaacg
     7681 aactggccct gccctccaaa tatgtgaact tcctgtacct ggccagccac tatgagaagc
     7741 tgaagggctc ccccgaggat aatgagcaga aacagctgtt tgtggaacag cacaaacact
     7801 acctggacga gatcatcgag cagatcagcg agttctccaa gagagtgatc ctggccgacg
     7861 ctaatctgga caaggtgctg agcgcctaca acaagcacag agacaagcct atcagagagc
     7921 aggccgagaa tatcatccac ctgtttaccc tgaccaatct gggagcccct gccgccttca
     7981 agtactttga caccaccatc gaccggaaga ggtacaccag caccaaagag gtgctggacg
     8041 ccaccctgat ccaccagagc atcaccggcc tgtacgagac acggatcgac ctgtctcagc
     8101 tgggaggcga cgcctatccc tatgacgtgc ccgattatgc cagcctgggc agcggctccc
     8161 ccaagaaaaa acgcaaggtg gaagatccta agaaaaagcg gaaagtggac ggcattggta
     8221 gtgggagcaa cggcagcagc ggatcctaac gcgtgaattc actcctcagg tgcaggctgc
     8281 ctatcagaag gtggtggctg gtgtggccaa tgccctggct cacaaatacc actgagatct
     8341 ttttccctct gccaaaaatt atggggacat catgaagccc cttgagcatc tgacttctgg
     8401 ctaataaagg aaatttattt tcattgcaat agtgtgttgg aattttttgt gtctctcact
     8461 cggaaggaca tatgggaggg caaatcattt aaaacatcag aatgagtatt tggtttagag
     8521 tttggcaaca tatgcccata tgctggctgc catgaacaaa ggttggctat aaagaggtca
     8581 tcagtatatg aaacagcccc ctgctgtcca ttccttattc catagaaaag ccttgacttg
     8641 aggttagatt ttttttatat tttgttttgt gttatttttt tctttaacat ccctaaaatt
     8701 ttccttacat gttttactag ccagattttt cctcctctcc tgactactcc cagtcatagc
     8761 tgtccctctt ctcttatgga gatccctcga cctgcagccc aagcttataa cttcgtatag
     8821 catacattat acgaagttat gcggccgcac cggtatgcat ttaattaaga tatcagcttt
     8881 actagggaca ggattggtga cagaaaagcc ccatccttag gcctcctcct tcctagtctc
     8941 ctgatattgg gtctaacccc cacctcctgt taggcagatt ccttatctgg tgacacaccc
     9001 ccatttcctg gagccatctc tctccttgcc agaacctcta aggtttgctt acgatggagc
     9061 cagagaggat cctgggaggg agagcttggc agggggtggg agggaagggg gggatgcgtg
     9121 acctgcccgg ttctcagtgg ccaccctgcg ctaccctctc ccagaacctg agctgctctg
     9181 acgcggctgt ctggtgcgtt tcactgatcc tggtgctgca gcttccttac acttcccaag
     9241 aggagaagca gtttggaaaa acaaaatcag aataagttgg tcctgagttc taactttggc
     9301 tcttcacctt tctagtcccc aatttatatt gttcctccgt gcgtcagttt tacctgtgag
     9361 ataaggccag tagccagccc cgtcctggca gggctgtggt gaggaggggg gtgtccgtgt
     9421 ggaaaactcc ctttgtgaga atggtgcgtc ctaggtgttc accaggtcgt ggccgcctct
     9481 actccctttc tctttctcca tccttctttc cttaaagagt ccccagtgct atctgggaca
     9541 tattcctccg cccagagcag ggtcccgctt ccctaaggcc ctgctctggg cttctgggtt
     9601 tgagtccttg gcaagcccag gagaggcgct caggcttccc tgtccccctt cctcgtccac
     9661 catctcatgc ccctggctct cctgcccctt ccctacaggg gttcctggct ctgctctaag
     9721 ggcgaattcg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     9781 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     9841 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     9901 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat
     9961 agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
    10021 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
    10081 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
    10141 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
    10201 agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
    10261 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
    10321 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
    10381 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
    10441 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
    10501 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
    10561 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
    10621 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
    10681 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
    10741 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
    10801 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
    10861 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
    10921 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
    10981 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
    11041 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
    11101 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
    11161 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
    11221 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
    11281 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
    11341 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
    11401 tattattgaa gcatttatca gggttgaatt cgccctt
//