LOCUS       sequence-244358-       14598 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express a dCas9, 10X GCN4, and mCherry cassette from the TRE3G 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14598)
  AUTHORS   Liu Y, Yu C, Daley TP, Wang F, Cao WS, Bhate S, Lin X, Still C 2nd, 
            Liu H, Zhao D, Wang H, Xie XS, Ding S, Wong WH, Wernig M, Qi LS
  TITLE     CRISPR Activation Screens Systematically Identify Factors that Drive
            Neuronal Fate and Reprogramming.
  JOURNAL   Cell Stem Cell. 2018 Sep 28. pii: S1934-5909(18)30440-5. doi: 
            10.1016/j.stem.2018.09.003.
  PUBMED    30318302
REFERENCE   2  (bases 1 to 14598)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cell Stem 
            Cell. 2018 Sep 28. pii: S1934-5909(18)30440-5. doi: 
            10.1016/j.stem.2018.09.003."
COMMENT     Sequence Label: pHR-TRE3G-dCas9-10X GCN4-P2A-mCherry
FEATURES             Location/Qualifiers
     source          1..14598
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(1..17)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             107..340
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     promoter        complement(365..383)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(366..383)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(673..692)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     complement(906..925)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1025..1047
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1085..1103)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1171..1275
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1276..1344,1345..2136)
                     /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     complement(1494..1513)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2307..2895
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2796..2815
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3049..3066
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     complement(3136..3156)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3141..3470
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3321..3456
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     intron          4604..4669
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     CDS             4799..4819
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     polyA_signal    5244..5378
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5281..5300)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5335..5354
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     LTR             5546..6179
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    6226..6351
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6848..7081
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             7266..7310
                     /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    7595..7712
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        7774..8123
                     /label=TRE3GV promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G, optimized for 
                     retroviral and lentiviral vectors"
     protein_bind    7782..7800
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7818..7836
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7854..7872
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7890..7908
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7926..7944
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7962..7980
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    7998..8016
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     8062..8086
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             join(8167..8193,8194..8196,8197..10683,10684..10686,
                     10687..12270)
                     /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             8167..12270
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /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             12274..12300
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             12280..12306
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     CDS             12319..12339
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             12346..12366
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             12442..12498
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             12514..12570
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             12586..12642
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             12658..12714
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             12730..12786
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             12802..12858
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             12874..12930
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             12946..13002
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             13018..13074
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             13090..13146
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             13180..13236
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             13237..13947
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(13382..13400)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     13650..13669
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    14007..14595
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(14060..14080)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(14478..14489)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(join(14598,1..16))
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 gataccgtcg acctcgaggg aattaattcg agctcggtac ctttaagacc aatgacttac
       61 aaggcagctg tagatcttag ccacttttta aaagaaaagg ggggactgga agggctaatt
      121 cactcccaac gaagacaaga tctgcttttt gcttgtactg ggtctctctg gttagaccag
      181 atctgagcct gggagctctc tggctaacta gggaacccac tgcttaagcc tcaataaagc
      241 ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg taactagaga
      301 tccctcagac ccttttagtc agtgtggaaa atctctagca gcatctagaa ttaattccgt
      361 gtattctata gtgtcaccta aatcgtatgt gtatgataca taaggttatg tattaattgt
      421 agccgcgttc taacgacaat atgtacaagc ctaattgtgt agcatctggc ttactgaagc
      481 agaccctatc atctctctcg taaactgccg tcagagtcgg tttggttgga cgaaccttct
      541 gagtttctgg taacgccgtc ccgcacccgg aaatggtcag cgaaccaatc agcagggtca
      601 tcgctagcca gatcctctac gccggacgca tcgtggccgg catcaccggc gccacaggtg
      661 cggttgctgg cgcctatatc gccgacatca ccgatgggga agatcgggct cgccacttcg
      721 ggctcatgag cgcttgtttc ggcgtgggta tggtggcagg ccccgtggcc gggggactgt
      781 tgggcgccat ctccttgcat gcaccattcc ttgcggcggc ggtgctcaac ggcctcaacc
      841 tactactggg ctgcttccta atgcaggagt cgcataaggg agagcgtcga atggtgcact
      901 ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc
      961 gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc
     1021 gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga
     1081 aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag
     1141 acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa
     1201 atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat
     1261 tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg
     1321 gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa
     1381 gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt
     1441 gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt
     1501 ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat
     1561 tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg
     1621 acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta
     1681 cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat
     1741 catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag
     1801 cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa
     1861 ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca
     1921 ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc
     1981 ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt
     2041 atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc
     2101 gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat
     2161 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     2221 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     2281 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     2341 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     2401 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     2461 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     2521 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     2581 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     2641 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     2701 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     2761 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     2821 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     2881 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     2941 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     3001 gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg
     3061 agcgaggaag cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt
     3121 cattaatgca gctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag
     3181 caggcagaag tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc
     3241 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag
     3301 tcccgcccct aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc
     3361 cccatggctg actaattttt tttatttatg cagaggccga ggccgcctcg gcctctgagc
     3421 tattccagaa gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagcttggac
     3481 acaagacagg cttgcgagat atgtttgaga ataccacttt atcccgcgtc agggagaggc
     3541 agtgcgtaaa aagacgcgga ctcatgtgaa atactggttt ttagtgcgcc agatctctat
     3601 aatctcgcgc aacctatttt cccctcgaac actttttaag ccgtagataa acaggctggg
     3661 acacttcaca tgagcgaaaa atacatcgtc acctgggaca tgttgcagat ccatgcacgt
     3721 aaactcgcaa gccgactgat gccttctgaa caatggaaag gcattattgc cgtaagccgt
     3781 ggcggtctgt accgggtgcg ttactggcgc gtgaactggg tattcgtcat gtcgataccg
     3841 tttgtatttc cagctacgat cacgacaacc agcgcgagct taaagtgctg aaacgcgcag
     3901 aaggcgatgg cgaaggcttc atcgttattg atgacctggt ggataccggt ggtactgcgg
     3961 ttgcgattcg tgaaatgtat ccaaaagcgc actttgtcac catcttcgca aaaccggctg
     4021 gtcgtccgct ggttgatgac tatgttgttg atatcccgca agatacctgg attgaacagc
     4081 cgtgggatat gggcgtcgta ttcgtcccgc caatctccgg tcgctaatct tttcaacgcc
     4141 tggcactgcc gggcgttgtt ctttttaact tcaggcgggt tacaatagtt tccagtaagt
     4201 attctggagg ctgcatccat gacacaggca aacctgagcg aaaccctgtt caaaccccgc
     4261 tttaaacatc ctgaaacctc gacgctagtc cgccgcttta atcacggcgc acaaccgcct
     4321 gtgcagtcgg cccttgatgg taaaaccatc cctcactggt atcgcatgat taaccgtctg
     4381 atgtggatct ggcgcggcat tgacccacgc gaaatcctcg acgtccaggc acgtattgtg
     4441 atgagcgatg ccgaacgtac cgacgatgat ttatacgata cggtgattgg ctaccgtggc
     4501 ggcaactgga tttatgagtg ggccccggat ctttgtgaag gaaccttact tctgtggtgt
     4561 gacataattg gacaaactac ctacagagat ttaaagctct aaggtaaata taaaattttt
     4621 aagtgtataa tgtgttaaac tactgattct aattgtttgt gtattttaga ttccaaccta
     4681 tggaactgat gaatgggagc agtggtggaa tgcctttaat gaggaaaacc tgttttgctc
     4741 agaagaaatg ccatctagtg atgatgaggc tactgctgac tctcaacatt ctactcctcc
     4801 aaaaaagaag agaaaggtag aagaccccaa ggactttcct tcagaattgc taagtttttt
     4861 gagtcatgct gtgtttagta atagaactct tgcttgcttt gctatttaca ccacaaagga
     4921 aaaagctgca ctgctataca agaaaattat ggaaaaatat tctgtaacct ttataagtag
     4981 gcataacagt tataatcata acatactgtt ttttcttact ccacacaggc atagagtgtc
     5041 tgctattaat aactatgctc aaaaattgtg tacctttagc tttttaattt gtaaaggggt
     5101 taataaggaa tatttgatgt atagtgcctt gactagagat cataatcagc cataccacat
     5161 ttgtagaggt tttacttgct ttaaaaaacc tcccacacct ccccctgaac ctgaaacata
     5221 aaatgaatgc aattgttgtt gttaacttgt ttattgcagc ttataatggt tacaaataaa
     5281 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     5341 tgtccaaact catcaatgta tcttatcatg tctggatcaa ctggataact caagctaacc
     5401 aaaatcatcc caaacttccc accccatacc ctattaccac tgccaattac ctgtggtttc
     5461 atttactcta aacctgtgat tcctctgaat tattttcatt ttaaagaaat tgtatttgtt
     5521 aaatatgtac tacaaactta gtagttggaa gggctaattc actcccaaag aagacaagat
     5581 atccttgatc tgtggatcta ccacacacaa ggctacttcc ctgattagca gaactacaca
     5641 ccagggccag gggtcagata tccactgacc tttggatggt gctacaagct agtaccagtt
     5701 gagccagata aggtagaaga ggccaataaa ggagagaaca ccagcttgtt acaccctgtg
     5761 agcctgcatg ggatggatga cccggagaga gaagtgttag agtggaggtt tgacagccgc
     5821 ctagcatttc atcacgtggc ccgagagctg catccggagt acttcaagaa ctgctgatat
     5881 cgagcttgct acaagggact ttccgctggg gactttccag ggaggcgtgg cctgggcggg
     5941 actggggagt ggcgagccct cagatcctgc atataagcag ctgctttttg cctgtactgg
     6001 gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact
     6061 gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg
     6121 tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag
     6181 tggcgcccga acagggactt gaaagcgaaa gggaaaccag aggagctctc tcgacgcagg
     6241 actcggcttg ctgaagcgcg cacggcaaga ggcgaggggc ggcgactggt gagtacgcca
     6301 aaaattttga ctagcggagg ctagaaggag agagatgggt gcgagagcgt cagtattaag
     6361 cgggggagaa ttagatcgcg atgggaaaaa attcggttaa ggccaggggg aaagaaaaaa
     6421 tataaattaa aacatatagt atgggcaagc agggagctag aacgattcgc agttaatcct
     6481 ggcctgttag aaacatcaga aggctgtaga caaatactgg gacagctaca accatccctt
     6541 cagacaggat cagaagaact tagatcatta tataatacag tagcaaccct ctattgtgtg
     6601 catcaaagga tagagataaa agacaccaag gaagctttag acaagataga ggaagagcaa
     6661 aacaaaagta agaccaccgc acagcaagcg gccggccgct gatcttcaga cctggaggag
     6721 gagatatgag ggacaattgg agaagtgaat tatataaata taaagtagta aaaattgaac
     6781 cattaggagt agcacccacc aaggcaaaga gaagagtggt gcagagagaa aaaagagcag
     6841 tgggaatagg agctttgttc cttgggttct tgggagcagc aggaagcact atgggcgcag
     6901 cgtcaatgac gctgacggta caggccagac aattattgtc tggtatagtg cagcagcaga
     6961 acaatttgct gagggctatt gaggcgcaac agcatctgtt gcaactcaca gtctggggca
     7021 tcaagcagct ccaggcaaga atcctggctg tggaaagata cctaaaggat caacagctcc
     7081 tggggatttg gggttgctct ggaaaactca tttgcaccac tgctgtgcct tggaatgcta
     7141 gttggagtaa taaatctctg gaacagattt ggaatcacac gacctggatg gagtgggaca
     7201 gagaaattaa caattacaca agcttaatac actccttaat tgaagaatcg caaaaccagc
     7261 aagaaaagaa tgaacaagaa ttattggaat tagataaatg ggcaagtttg tggaattggt
     7321 ttaacataac aaattggctg tggtatataa aattattcat aatgatagta ggaggcttgg
     7381 taggtttaag aatagttttt gctgtacttt ctatagtgaa tagagttagg cagggatatt
     7441 caccattatc gtttcagacc cacctcccaa ccccgagggg acccgacagg cccgaaggaa
     7501 tagaagaaga aggtggagag agagacagag acagatccat tcgattagtg aacggatctc
     7561 gacggtatcg ccaaatggca gtattcatcc acaattttaa aagaaaaggg gggattgggg
     7621 ggtacagtgc aggggaaaga atagtagaca taatagcaac agacatacaa actaaagaat
     7681 tacaaaaaca aattacaaaa attcaaaatt ttcgggttta ttacagggac agcagagatc
     7741 cagtttatcg atgaggccct ttcgtcttca ctcgagttta ctccctatca gtgatagaga
     7801 acgtatgaag agtttactcc ctatcagtga tagagaacgt atgcagactt tactccctat
     7861 cagtgataga gaacgtataa ggagtttact ccctatcagt gatagagaac gtatgaccag
     7921 tttactccct atcagtgata gagaacgtat ctacagttta ctccctatca gtgatagaga
     7981 acgtatatcc agtttactcc ctatcagtga tagagaacgt atgtcgaggt aggcgtgtac
     8041 ggtgggcgcc tataaaagca gagctcgttt agtgaaccgt cagatcgcct ggagcaattc
     8101 cacaacactt ttgtcttata cttacgcgtc tgcaggatat caagcttgcg gtaccgcggg
     8161 cccgggatgg acaagaagta cagcatcggc ctggccatcg gcaccaactc tgtgggctgg
     8221 gccgtgatca ccgacgagta caaggtgccc agcaagaaat tcaaggtgct gggcaacacc
     8281 gaccggcaca gcatcaagaa gaacctgatc ggcgccctgc tgttcgacag cggagaaaca
     8341 gccgaggcca cccggctgaa gagaaccgcc agaagaagat acaccagacg gaagaaccgg
     8401 atctgctatc tgcaagagat cttcagcaac gagatggcca aggtggacga cagcttcttc
     8461 cacagactgg aagagtcctt cctggtggaa gaggataaga agcacgagcg gcaccccatc
     8521 ttcggcaaca tcgtggacga ggtggcctac cacgagaagt accccaccat ctaccacctg
     8581 agaaagaaac tggtggacag caccgacaag gccgacctgc ggctgatcta tctggccctg
     8641 gcccacatga tcaagttccg gggccacttc ctgatcgagg gcgacctgaa ccccgacaac
     8701 agcgacgtgg acaagctgtt catccagctg gtgcagacct acaaccagct gttcgaggaa
     8761 aaccccatca acgccagcgg cgtggacgcc aaggccatcc tgtctgccag actgagcaag
     8821 agcagacggc tggaaaatct gatcgcccag ctgcccggcg agaagaagaa tggcctgttc
     8881 ggcaacctga ttgccctgag cctgggcctg acccccaact tcaagagcaa cttcgacctg
     8941 gccgaggatg ccaaactgca gctgagcaag gacacctacg acgacgacct ggacaacctg
     9001 ctggcccaga tcggcgacca gtacgccgac ctgtttctgg ccgccaagaa cctgtccgac
     9061 gccatcctgc tgagcgacat cctgagagtg aacaccgaga tcaccaaggc ccccctgagc
     9121 gcctctatga tcaagagata cgacgagcac caccaggacc tgaccctgct gaaagctctc
     9181 gtgcggcagc agctgcctga gaagtacaaa gagattttct tcgaccagag caagaacggc
     9241 tacgccggct acatcgatgg cggagccagc caggaagagt tctacaagtt catcaagccc
     9301 atcctggaaa agatggacgg caccgaggaa ctgctcgtga agctgaacag agaggacctg
     9361 ctgcggaagc agcggacctt cgacaacggc agcatccccc accagatcca cctgggagag
     9421 ctgcacgcca ttctgcggcg gcaggaagat ttttacccat tcctgaagga caaccgggaa
     9481 aagatcgaga agatcctgac cttccgcatc ccctactacg tgggccctct ggccagggga
     9541 aacagcagat tcgcctggat gaccagaaag agcgaggaaa ccatcacccc ctggaacttc
     9601 gaggaagtgg tggacaaggg cgccagcgcc cagagcttca tcgagcggat gaccaacttc
     9661 gataagaacc tgcccaacga gaaggtgctg cccaagcaca gcctgctgta cgagtacttc
     9721 accgtgtaca acgagctgac caaagtgaaa tacgtgaccg agggaatgag aaagcccgcc
     9781 ttcctgagcg gcgagcagaa aaaagccatc gtggacctgc tgttcaagac caaccggaaa
     9841 gtgaccgtga agcagctgaa agaggactac ttcaagaaaa tcgagtgctt cgactccgtg
     9901 gaaatctccg gcgtggaaga tcggttcaac gcctccctgg gcacatacca cgatctgctg
     9961 aaaattatca aggacaagga cttcctggac aatgaggaaa acgaggacat tctggaagat
    10021 atcgtgctga ccctgacact gtttgaggac agagagatga tcgaggaacg gctgaaaacc
    10081 tatgcccacc tgttcgacga caaagtgatg aagcagctga agcggcggag atacaccggc
    10141 tggggcaggc tgagccggaa gctgatcaac ggcatccggg acaagcagtc cggcaagaca
    10201 atcctggatt tcctgaagtc cgacggcttc gccaacagaa acttcatgca gctgatccac
    10261 gacgacagcc tgacctttaa agaggacatc cagaaagccc aggtgtccgg ccagggcgat
    10321 agcctgcacg agcacattgc caatctggcc ggcagccccg ccattaagaa gggcatcctg
    10381 cagacagtga aggtggtgga cgagctcgtg aaagtgatgg gccggcacaa gcccgagaac
    10441 atcgtgatcg aaatggccag agagaaccag accacccaga agggacagaa gaacagccgc
    10501 gagagaatga agcggatcga agagggcatc aaagagctgg gcagccagat cctgaaagaa
    10561 caccccgtgg aaaacaccca gctgcagaac gagaagctgt acctgtacta cctgcagaat
    10621 gggcgggata tgtacgtgga ccaggaactg gacatcaacc ggctgtccga ctacgatgtg
    10681 gacgctatcg tgcctcagag ctttctgaag gacgactcca tcgataacaa agtgctgact
    10741 cggagcgaca agaaccgggg caagagcgac aacgtgccct ccgaagaggt cgtgaagaag
    10801 atgaagaact actggcgcca gctgctgaat gccaagctga ttacccagag gaagttcgac
    10861 aatctgacca aggccgagag aggcggcctg agcgaactgg ataaggccgg cttcatcaag
    10921 agacagctgg tggaaacccg gcagatcaca aagcacgtgg cacagatcct ggactcccgg
    10981 atgaacacta agtacgacga gaacgacaaa ctgatccggg aagtgaaagt gatcaccctg
    11041 aagtccaagc tggtgtccga tttccggaag gatttccagt tttacaaagt gcgcgagatc
    11101 aacaactacc accacgccca cgacgcctac ctgaacgccg tcgtgggaac cgccctgatc
    11161 aaaaagtacc ctaagctgga aagcgagttc gtgtacggcg actacaaggt gtacgacgtg
    11221 cggaagatga tcgccaagag cgagcaggaa atcggcaagg ctaccgccaa gtacttcttc
    11281 tacagcaaca tcatgaactt tttcaagacc gagattaccc tggccaacgg cgagatccgg
    11341 aagcggcctc tgatcgagac aaacggcgaa acaggcgaga tcgtgtggga taagggccgg
    11401 gactttgcca ccgtgcggaa agtgctgtct atgccccaag tgaatatcgt gaaaaagacc
    11461 gaggtgcaga caggcggctt cagcaaagag tctatcctgc ccaagaggaa cagcgacaag
    11521 ctgatcgcca gaaagaagga ctgggaccct aagaagtacg gcggcttcga cagccccacc
    11581 gtggcctatt ctgtgctggt ggtggccaaa gtggaaaagg gcaagtccaa gaaactgaag
    11641 agtgtgaaag agctgctggg gatcaccatc atggaaagaa gcagcttcga gaagaatccc
    11701 atcgactttc tggaagccaa gggctacaaa gaagtgaaaa aggacctgat catcaagctg
    11761 cctaagtact ccctgttcga gctggaaaac ggccggaaga gaatgctggc ctctgccggc
    11821 gaactgcaga agggaaacga actggccctg ccctccaaat atgtgaactt cctgtacctg
    11881 gccagccact atgagaagct gaagggctcc cccgaggata atgagcagaa acagctgttt
    11941 gtggaacagc acaaacacta cctggacgag atcatcgagc agatcagcga gttctccaag
    12001 agagtgatcc tggccgacgc taatctggac aaggtgctga gcgcctacaa caagcacaga
    12061 gacaagccta tcagagagca ggccgagaat atcatccacc tgtttaccct gaccaatctg
    12121 ggagcccctg ccgccttcaa gtactttgac accaccatcg accggaagag gtacaccagc
    12181 accaaagagg tgctggacgc caccctgatc caccagagca tcaccggcct gtacgagaca
    12241 cggatcgacc tgtctcagct gggaggcgac gcctatccct atgacgtgcc cgattatgcc
    12301 agcctgggca gcggctcccc caagaaaaaa cgcaaggtgg aagatcctaa gaaaaagcgg
    12361 aaagtggacg gcattggtag tgggagcaac ggcagcagcg gatccggcgc catcatacac
    12421 gcggccgcaa caagcggtcc ggaagaactt ttgagcaaga attatcatct tgagaacgaa
    12481 gtggctcgtc ttaagaaagg ttctggcagt ggagaagaac tgctttcaaa gaattaccac
    12541 ctggaaaatg aggtagctag actgaaaaag gggagcggaa gtggggagga gttgctgagc
    12601 aaaaattatc atttggagaa cgaagtagca cgactaaaga aagggtccgg atcgggtgag
    12661 gagttactct cgaaaaatta tcatctcgaa aacgaagtgg ctcggctaaa aaagggcagt
    12721 ggttctggag aagagctatt atctaaaaac taccacctcg aaaatgaggt ggcacgctta
    12781 aaaaagggaa gtggcagtgg tgaagagcta ctatccaaga attatcatct tgagaacgag
    12841 gtagcgcgtt tgaagaaggg ttccggctca ggagaggaac tgctctcgaa gaactatcat
    12901 cttgaaaatg aggtcgctcg attaaaaaag ggatcgggca gtggtgagga actactttca
    12961 aagaattacc acctcgaaaa cgaagtagct cgattaaaga aaggttcagg gtcgggtgaa
    13021 gaattactga gtaaaaatta tcatctggaa aatgaggtag cgagactaaa aaaggggagt
    13081 ggttctggcg aggaattgct atcgaaaaat tatcatcttg agaacgaagt tgctaggctc
    13141 aaaaagggct caggctcagg caaaggtccg ggaagcggag ctactaactt cagcctgctg
    13201 aagcaggctg gagacgtgga ggagaaccct ggacctatgg tgagcaaggg cgaggaggat
    13261 aacatggcca tcatcaagga gttcatgcgc ttcaaggtgc acatggaggg ctccgtgaac
    13321 ggccacgagt tcgagatcga gggcgagggc gagggccgcc cctacgaggg cacccagacc
    13381 gccaagctga aggtgaccaa gggtggcccc ctgcccttcg cctgggacat cctgtcccct
    13441 cagttcatgt acggctccaa ggcctacgtg aagcaccccg ccgacatccc cgactacttg
    13501 aagctgtcct tccccgaggg cttcaagtgg gagcgcgtga tgaacttcga ggacggcggc
    13561 gtggtgaccg tgacccagga ctcctccctg caggacggcg agttcatcta caaggtgaag
    13621 ctgcgcggca ccaacttccc ctccgacggc cccgtaatgc agaagaagac catgggctgg
    13681 gaggcctcct ccgagcggat gtaccccgag gacggcgccc tgaagggcga gatcaagcag
    13741 aggctgaagc tgaaggacgg cggccactac gacgctgagg tcaagaccac ctacaaggcc
    13801 aagaagcccg tgcagctgcc cggcgcctac aacgtcaaca tcaagttgga catcacctcc
    13861 cacaacgagg actacaccat cgtggaacag tacgaacgcg ccgagggccg ccactccacc
    13921 ggcggcatgg acgagctgta caagtaagcg gccgcgactc tagagtcgac ctgcaggcat
    13981 gcaagcttga tatcaagctt atcgataatc aacctctgga ttacaaaatt tgtgaaagat
    14041 tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc
    14101 ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct
    14161 ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca
    14221 ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt
    14281 ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg
    14341 cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga
    14401 aatcatcgtc ctttccttgg ctgctcgcct gtgttgccac ctggattctg cgcgggacgt
    14461 ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc
    14521 cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt
    14581 gggccgcctc cccgcatc
//
