LOCUS       sequence-180844-       13013 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  sgNf1.1 for Nf1 deletion.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13013)
  AUTHORS   Song CQ, Li Y, Mou H, Moore J, Park A, Pomyen Y, Hough S, Kennedy Z,
            Fischer A, Yin H, Anderson DG, Conte D Jr, Zender L, Wang XW, 
            Thorgeirsson S, Weng Z, Xue W
  TITLE     Genome-Wide CRISPR Screen Identifies Regulators of Mitogen-Activated
            Protein Kinase as Suppressors of Liver Tumors in Mice.
  JOURNAL   Gastroenterology. 2017 Apr;152(5):1161-1173.e1. doi: 
            10.1053/j.gastro.2016.12.002. Epub 2016 Dec 10.
  PUBMED    27956228
REFERENCE   2  (bases 1 to 13013)
  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"; doi: 
            "10.1053/j.gastro.2016.12.002"; journalName: "Gastroenterology"; 
            date: "2017-04"; volume: "152"; issue: "5"; pages: "1161-1173.e1"
COMMENT     Sequence Label: lentiCRISPRv2.sgNf1.1
FEATURES             Location/Qualifiers
     source          1..13013
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..249
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1..21
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     172..191
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        271..346
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        408..619
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      638..647
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             644..4747
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             644..4747
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             4748..4795
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             4796..4819
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             4829..4885
                     /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             4886..5482
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
                     TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
                     QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
                     PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     5379..5399
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    5498..6086
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5551..5571)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5969..5980)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             6158..6391
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(6417..6434)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6423..6647
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      6693..7121
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6780..6799)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6990..7011
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(7130..7150)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        7135..7464
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7315..7450
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7377..7396
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7512..7559
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7578..7952
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    8082..8203
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8119..8138)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8173..8192
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(8252..8268)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8252..8268)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(8265..8287)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    8276..8292
                     /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(join(8300..8306,8307..8324,8325..8330))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8345..8366
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(8483..8500)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8654..9242)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8734..8753)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(9413..10204,10205..10273))
                     /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     10036..10055
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(10274..10378)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(10453..10472)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        10644..11023
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        11025..11223
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     11178..11198
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             11241..11421
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    11468..11593
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    12086..12319
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             12504..12548
                     /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    12846..12963
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
       61 ataattagaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
      121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
      181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
      241 cgaaacaccg tccgaagttc ggctgcatgt gttttagagc tagaaatagc aagttaaaat
      301 aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt ttgaattcgc
      361 tagctaggtc ttgaaaggag tgggaattgg ctccggtgcc cgtcagtggg cagagcgcac
      421 atcgcccaca gtccccgaga agttgggggg aggggtcggc aattgatccg gtgcctagag
      481 aaggtggcgc ggggtaaact gggaaagtga tgtcgtgtac tggctccgcc tttttcccga
      541 gggtggggga gaaccgtata taagtgcagt agtcgccgtg aacgttcttt ttcgcaacgg
      601 gtttgccgcc agaacacagg accggttcta gagcgctgcc accatggaca agaagtacag
      661 catcggcctg gacatcggca ccaactctgt gggctgggcc gtgatcaccg acgagtacaa
      721 ggtgcccagc aagaaattca aggtgctggg caacaccgac cggcacagca tcaagaagaa
      781 cctgatcgga gccctgctgt tcgacagcgg cgaaacagcc gaggccaccc ggctgaagag
      841 aaccgccaga agaagataca ccagacggaa gaaccggatc tgctatctgc aagagatctt
      901 cagcaacgag atggccaagg tggacgacag cttcttccac agactggaag agtccttcct
      961 ggtggaagag gataagaagc acgagcggca ccccatcttc ggcaacatcg tggacgaggt
     1021 ggcctaccac gagaagtacc ccaccatcta ccacctgaga aagaaactgg tggacagcac
     1081 cgacaaggcc gacctgcggc tgatctatct ggccctggcc cacatgatca agttccgggg
     1141 ccacttcctg atcgagggcg acctgaaccc cgacaacagc gacgtggaca agctgttcat
     1201 ccagctggtg cagacctaca accagctgtt cgaggaaaac cccatcaacg ccagcggcgt
     1261 ggacgccaag gccatcctgt ctgccagact gagcaagagc agacggctgg aaaatctgat
     1321 cgcccagctg cccggcgaga agaagaatgg cctgttcgga aacctgattg ccctgagcct
     1381 gggcctgacc cccaacttca agagcaactt cgacctggcc gaggatgcca aactgcagct
     1441 gagcaaggac acctacgacg acgacctgga caacctgctg gcccagatcg gcgaccagta
     1501 cgccgacctg tttctggccg ccaagaacct gtccgacgcc atcctgctga gcgacatcct
     1561 gagagtgaac accgagatca ccaaggcccc cctgagcgcc tctatgatca agagatacga
     1621 cgagcaccac caggacctga ccctgctgaa agctctcgtg cggcagcagc tgcctgagaa
     1681 gtacaaagag attttcttcg accagagcaa gaacggctac gccggctaca ttgacggcgg
     1741 agccagccag gaagagttct acaagttcat caagcccatc ctggaaaaga tggacggcac
     1801 cgaggaactg ctcgtgaagc tgaacagaga ggacctgctg cggaagcagc ggaccttcga
     1861 caacggcagc atcccccacc agatccacct gggagagctg cacgccattc tgcggcggca
     1921 ggaagatttt tacccattcc tgaaggacaa ccgggaaaag atcgagaaga tcctgacctt
     1981 ccgcatcccc tactacgtgg gccctctggc caggggaaac agcagattcg cctggatgac
     2041 cagaaagagc gaggaaacca tcaccccctg gaacttcgag gaagtggtgg acaagggcgc
     2101 ttccgcccag agcttcatcg agcggatgac caacttcgat aagaacctgc ccaacgagaa
     2161 ggtgctgccc aagcacagcc tgctgtacga gtacttcacc gtgtataacg agctgaccaa
     2221 agtgaaatac gtgaccgagg gaatgagaaa gcccgccttc ctgagcggcg agcagaaaaa
     2281 ggccatcgtg gacctgctgt tcaagaccaa ccggaaagtg accgtgaagc agctgaaaga
     2341 ggactacttc aagaaaatcg agtgcttcga ctccgtggaa atctccggcg tggaagatcg
     2401 gttcaacgcc tccctgggca cataccacga tctgctgaaa attatcaagg acaaggactt
     2461 cctggacaat gaggaaaacg aggacattct ggaagatatc gtgctgaccc tgacactgtt
     2521 tgaggacaga gagatgatcg aggaacggct gaaaacctat gcccacctgt tcgacgacaa
     2581 agtgatgaag cagctgaagc ggcggagata caccggctgg ggcaggctga gccggaagct
     2641 gatcaacggc atccgggaca agcagtccgg caagacaatc ctggatttcc tgaagtccga
     2701 cggcttcgcc aacagaaact tcatgcagct gatccacgac gacagcctga cctttaaaga
     2761 ggacatccag aaagcccagg tgtccggcca gggcgatagc ctgcacgagc acattgccaa
     2821 tctggccggc agccccgcca ttaagaaggg catcctgcag acagtgaagg tggtggacga
     2881 gctcgtgaaa gtgatgggcc ggcacaagcc cgagaacatc gtgatcgaaa tggccagaga
     2941 gaaccagacc acccagaagg gacagaagaa cagccgcgag agaatgaagc ggatcgaaga
     3001 gggcatcaaa gagctgggca gccagatcct gaaagaacac cccgtggaaa acacccagct
     3061 gcagaacgag aagctgtacc tgtactacct gcagaatggg cgggatatgt acgtggacca
     3121 ggaactggac atcaaccggc tgtccgacta cgatgtggac catatcgtgc ctcagagctt
     3181 tctgaaggac gactccatcg acaacaaggt gctgaccaga agcgacaaga accggggcaa
     3241 gagcgacaac gtgccctccg aagaggtcgt gaagaagatg aagaactact ggcggcagct
     3301 gctgaacgcc aagctgatta cccagagaaa gttcgacaat ctgaccaagg ccgagagagg
     3361 cggcctgagc gaactggata aggccggctt catcaagaga cagctggtgg aaacccggca
     3421 gatcacaaag cacgtggcac agatcctgga ctcccggatg aacactaagt acgacgagaa
     3481 tgacaagctg atccgggaag tgaaagtgat caccctgaag tccaagctgg tgtccgattt
     3541 ccggaaggat ttccagtttt acaaagtgcg cgagatcaac aactaccacc acgcccacga
     3601 cgcctacctg aacgccgtcg tgggaaccgc cctgatcaaa aagtacccta agctggaaag
     3661 cgagttcgtg tacggcgact acaaggtgta cgacgtgcgg aagatgatcg ccaagagcga
     3721 gcaggaaatc ggcaaggcta ccgccaagta cttcttctac agcaacatca tgaacttttt
     3781 caagaccgag attaccctgg ccaacggcga gatccggaag cggcctctga tcgagacaaa
     3841 cggcgaaacc ggggagatcg tgtgggataa gggccgggat tttgccaccg tgcggaaagt
     3901 gctgagcatg ccccaagtga atatcgtgaa aaagaccgag gtgcagacag gcggcttcag
     3961 caaagagtct atcctgccca agaggaacag cgataagctg atcgccagaa agaaggactg
     4021 ggaccctaag aagtacggcg gcttcgacag ccccaccgtg gcctattctg tgctggtggt
     4081 ggccaaagtg gaaaagggca agtccaagaa actgaagagt gtgaaagagc tgctggggat
     4141 caccatcatg gaaagaagca gcttcgagaa gaatcccatc gactttctgg aagccaaggg
     4201 ctacaaagaa gtgaaaaagg acctgatcat caagctgcct aagtactccc tgttcgagct
     4261 ggaaaacggc cggaagagaa tgctggcctc tgccggcgaa ctgcagaagg gaaacgaact
     4321 ggccctgccc tccaaatatg tgaacttcct gtacctggcc agccactatg agaagctgaa
     4381 gggctccccc gaggataatg agcagaaaca gctgtttgtg gaacagcaca agcactacct
     4441 ggacgagatc atcgagcaga tcagcgagtt ctccaagaga gtgatcctgg ccgacgctaa
     4501 tctggacaaa gtgctgtccg cctacaacaa gcaccgggat aagcccatca gagagcaggc
     4561 cgagaatatc atccacctgt ttaccctgac caatctggga gcccctgccg ccttcaagta
     4621 ctttgacacc accatcgacc ggaagaggta caccagcacc aaagaggtgc tggacgccac
     4681 cctgatccac cagagcatca ccggcctgta cgagacacgg atcgacctgt ctcagctggg
     4741 aggcgacaag cgacctgccg ccacaaagaa ggctggacag gctaagaaga agaaagatta
     4801 caaagacgat gacgataagg gatccggcgc aacaaacttc tctctgctga aacaagccgg
     4861 agatgtcgaa gagaatcctg gaccgaccga gtacaagccc acggtgcgcc tcgccacccg
     4921 cgacgacgtc cccagggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac
     4981 gcgccacacc gtcgatccgg accgccacat cgagcgggtc accgagctgc aagaactctt
     5041 cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt
     5101 ggcggtctgg accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc
     5161 gcgcatggcc gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct
     5221 ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc accgtcggag tctcgcccga
     5281 ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg
     5341 cgccggggtg cccgccttcc tggagacctc cgcgccccgc aacctcccct tctacgagcg
     5401 gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat
     5461 gacccgcaag cccggtgcct gaacgcgtta agtcgacaat caacctctgg attacaaaat
     5521 ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc
     5581 tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt
     5641 gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg
     5701 cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg
     5761 tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc
     5821 cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt
     5881 gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc tgtgttgcca cctggattct
     5941 gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg
     6001 cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg
     6061 gatctccctt tgggccgcct ccccgcgtcg actttaagac caatgactta caaggcagct
     6121 gtagatctta gccacttttt aaaagaaaag gggggactgg aagggctaat tcactcccaa
     6181 cgaagacaag atctgctttt tgcttgtact gggtctctct ggttagacca gatctgagcc
     6241 tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag cttgccttga
     6301 gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag atccctcaga
     6361 cccttttagt cagtgtggaa aatctctagc agggcccgtt taaacccgct gatcagcctc
     6421 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac
     6481 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg
     6541 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga
     6601 ttgggaagac aatagcaggc atgctgggga tgcggtgggc tctatggctt ctgaggcgga
     6661 aagaaccagc tggggctcta gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc
     6721 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     6781 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     6841 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     6901 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     6961 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     7021 caaccctatc tcggtctatt cttttgattt ataagggatt ttgccgattt cggcctattg
     7081 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt
     7141 cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     7201 ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     7261 caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg
     7321 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     7381 tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg aggaggcttt
     7441 tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat tttcggatct
     7501 gatcagcacg tgttgacaat taatcatcgg catagtatat cggcatagta taatacgaca
     7561 aggtgaggaa ctaaaccatg gccaagttga ccagtgccgt tccggtgctc accgcgcgcg
     7621 acgtcgccgg agcggtcgag ttctggaccg accggctcgg gttctcccgg gacttcgtgg
     7681 aggacgactt cgccggtgtg gtccgggacg acgtgaccct gttcatcagc gcggtccagg
     7741 accaggtggt gccggacaac accctggcct gggtgtgggt gcgcggcctg gacgagctgt
     7801 acgccgagtg gtcggaggtc gtgtccacga acttccggga cgcctccggg ccggccatga
     7861 ccgagatcgg cgagcagccg tgggggcggg agttcgccct gcgcgacccg gccggcaact
     7921 gcgtgcactt cgtggccgag gagcaggact gacacgtgct acgagatttc gattccaccg
     7981 ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc tggatgatcc
     8041 tccagcgcgg ggatctcatg ctggagttct tcgcccaccc caacttgttt attgcagctt
     8101 ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac
     8161 tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tgtataccgt
     8221 cgacctctag ctagagcttg gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     8281 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     8341 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     8401 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     8461 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     8521 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     8581 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     8641 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     8701 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     8761 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     8821 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     8881 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     8941 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     9001 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     9061 tgaagtggtg gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc
     9121 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     9181 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     9241 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     9301 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     9361 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     9421 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     9481 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     9541 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     9601 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     9661 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     9721 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     9781 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     9841 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     9901 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     9961 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
    10021 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
    10081 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
    10141 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
    10201 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
    10261 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
    10321 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
    10381 catttccccg aaaagtgcca cctgacgtcg acggatcggg agatctcccg atcccctatg
    10441 gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagtatc tgctccctgc
    10501 ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt taagctacaa caaggcaagg
    10561 cttgaccgac aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga
    10621 tgtacgggcc agatatacgc gttgacattg attattgact agttattaat agtaatcaat
    10681 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
    10741 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
    10801 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
    10861 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
    10921 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
    10981 tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca
    11041 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat
    11101 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
    11161 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
    11221 cagcgcgttt tgcctgtact gggtctctct ggttagacca gatctgagcc tgggagctct
    11281 ctggctaact agggaaccca ctgcttaagc ctcaataaag cttgccttga gtgcttcaag
    11341 tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag atccctcaga cccttttagt
    11401 cagtgtggaa aatctctagc agtggcgccc gaacagggac ttgaaagcga aagggaaacc
    11461 agaggagctc tctcgacgca ggactcggct tgctgaagcg cgcacggcaa gaggcgaggg
    11521 gcggcgactg gtgagtacgc caaaaatttt gactagcgga ggctagaagg agagagatgg
    11581 gtgcgagagc gtcagtatta agcgggggag aattagatcg cgatgggaaa aaattcggtt
    11641 aaggccaggg ggaaagaaaa aatataaatt aaaacatata gtatgggcaa gcagggagct
    11701 agaacgattc gcagttaatc ctggcctgtt agaaacatca gaaggctgta gacaaatact
    11761 gggacagcta caaccatccc ttcagacagg atcagaagaa cttagatcat tatataatac
    11821 agtagcaacc ctctattgtg tgcatcaaag gatagagata aaagacacca aggaagcttt
    11881 agacaagata gaggaagagc aaaacaaaag taagaccacc gcacagcaag cggccgctga
    11941 tcttcagacc tggaggagga gatatgaggg acaattggag aagtgaatta tataaatata
    12001 aagtagtaaa aattgaacca ttaggagtag cacccaccaa ggcaaagaga agagtggtgc
    12061 agagagaaaa aagagcagtg ggaataggag ctttgttcct tgggttcttg ggagcagcag
    12121 gaagcactat gggcgcagcg tcaatgacgc tgacggtaca ggccagacaa ttattgtctg
    12181 gtatagtgca gcagcagaac aatttgctga gggctattga ggcgcaacag catctgttgc
    12241 aactcacagt ctggggcatc aagcagctcc aggcaagaat cctggctgtg gaaagatacc
    12301 taaaggatca acagctcctg gggatttggg gttgctctgg aaaactcatt tgcaccactg
    12361 ctgtgccttg gaatgctagt tggagtaata aatctctgga acagatttgg aatcacacga
    12421 cctggatgga gtgggacaga gaaattaaca attacacaag cttaatacac tccttaattg
    12481 aagaatcgca aaaccagcaa gaaaagaatg aacaagaatt attggaatta gataaatggg
    12541 caagtttgtg gaattggttt aacataacaa attggctgtg gtatataaaa ttattcataa
    12601 tgatagtagg aggcttggta ggtttaagaa tagtttttgc tgtactttct atagtgaata
    12661 gagttaggca gggatattca ccattatcgt ttcagaccca cctcccaacc ccgaggggac
    12721 ccgacaggcc cgaaggaata gaagaagaag gtggagagag agacagagac agatccattc
    12781 gattagtgaa cggatcggca ctgcgtgcgc caattctgca gacaaatggc agtattcatc
    12841 cacaatttta aaagaaaagg ggggattggg gggtacagtg caggggaaag aatagtagac
    12901 ataatagcaa cagacataca aactaaagaa ttacaaaaac aaattacaaa aattcaaaat
    12961 tttcgggttt attacaggga cagcagagat ccagtttggt taattaaggt acc
//
