LOCUS       sequence-270138-       13400 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  3rd generation lentiviral gRNA plasmid targeting human CDKN1A.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13400)
  AUTHORS   Kwan SY, Sheel A, Song CQ, Zhang XO, Jiang T, Dang H, Cao Y, Ozata 
            DM, Mou H, Yin H, Weng Z, Wang XW, Xue W
  TITLE     Depletion of TRRAP induces p53-independent senescence in liver 
            cancer by downregulating mitotic genes.
  JOURNAL   Hepatology. 2019 Jun 12. doi: 10.1002/hep.30807.
  PUBMED    31188495
REFERENCE   2  (bases 1 to 13400)
  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: "Hepatology.
            2019 Jun 12. doi: 10.1002/hep.30807."
COMMENT     Sequence Label: CDKN1A sgRNA
FEATURES             Location/Qualifiers
     source          1..13400
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(47..66)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        242..621
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        623..821
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     776..796
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             839..1019
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1066..1191
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1684..1917
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2102..2146
                     /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    2444..2561
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2612..2860
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2612..2632
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2783..2802
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2881..2956
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        3018..3229
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      3248..3257
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3254..7357
                     /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             3254..7357
                     /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             7358..7405
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7406..7429
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             7439..7495
                     /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"
     misc_feature    8492..9080
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8545..8565)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8963..8974)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             9152..9385
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(9411..9428)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9417..9641
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      9687..10115
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9774..9793)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9984..10005
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10124..10144)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10129..10458
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      10309..10444
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10371..10390
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10506..10553
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10572..10946
                     /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    11076..11197
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11113..11132)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11167..11186
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11246..11262)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11246..11262)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11259..11281)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11270..11286
                     /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(11294..11300,11301..11318,11319..11324))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11339..11360
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11477..11494)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11648..12236)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11728..11747)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12407..13198,13199..13267))
                     /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     13030..13049
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(13268..13372)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgcgcgt
      241 tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt agttcatagc
      301 ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc
      361 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg
      421 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat
      481 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc
      541 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta
      601 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag
      661 cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt
      721 tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa
      781 atgggcggta ggcgtgtacg gtgggaggtc tatataagca gcgcgttttg cctgtactgg
      841 gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact
      901 gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg
      961 tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag
     1021 tggcgcccga acagggactt gaaagcgaaa gggaaaccag aggagctctc tcgacgcagg
     1081 actcggcttg ctgaagcgcg cacggcaaga ggcgaggggc ggcgactggt gagtacgcca
     1141 aaaattttga ctagcggagg ctagaaggag agagatgggt gcgagagcgt cagtattaag
     1201 cgggggagaa ttagatcgcg atgggaaaaa attcggttaa ggccaggggg aaagaaaaaa
     1261 tataaattaa aacatatagt atgggcaagc agggagctag aacgattcgc agttaatcct
     1321 ggcctgttag aaacatcaga aggctgtaga caaatactgg gacagctaca accatccctt
     1381 cagacaggat cagaagaact tagatcatta tataatacag tagcaaccct ctattgtgtg
     1441 catcaaagga tagagataaa agacaccaag gaagctttag acaagataga ggaagagcaa
     1501 aacaaaagta agaccaccgc acagcaagcg gccgctgatc ttcagacctg gaggaggaga
     1561 tatgagggac aattggagaa gtgaattata taaatataaa gtagtaaaaa ttgaaccatt
     1621 aggagtagca cccaccaagg caaagagaag agtggtgcag agagaaaaaa gagcagtggg
     1681 aataggagct ttgttccttg ggttcttggg agcagcagga agcactatgg gcgcagcgtc
     1741 aatgacgctg acggtacagg ccagacaatt attgtctggt atagtgcagc agcagaacaa
     1801 tttgctgagg gctattgagg cgcaacagca tctgttgcaa ctcacagtct ggggcatcaa
     1861 gcagctccag gcaagaatcc tggctgtgga aagataccta aaggatcaac agctcctggg
     1921 gatttggggt tgctctggaa aactcatttg caccactgct gtgccttgga atgctagttg
     1981 gagtaataaa tctctggaac agatttggaa tcacacgacc tggatggagt gggacagaga
     2041 aattaacaat tacacaagct taatacactc cttaattgaa gaatcgcaaa accagcaaga
     2101 aaagaatgaa caagaattat tggaattaga taaatgggca agtttgtgga attggtttaa
     2161 cataacaaat tggctgtggt atataaaatt attcataatg atagtaggag gcttggtagg
     2221 tttaagaata gtttttgctg tactttctat agtgaataga gttaggcagg gatattcacc
     2281 attatcgttt cagacccacc tcccaacccc gaggggaccc gacaggcccg aaggaataga
     2341 agaagaaggt ggagagagag acagagacag atccattcga ttagtgaacg gatcggcact
     2401 gcgtgcgcca attctgcaga caaatggcag tattcatcca caattttaaa agaaaagggg
     2461 ggattggggg gtacagtgca ggggaaagaa tagtagacat aatagcaaca gacatacaaa
     2521 ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt tcgggtttat tacagggaca
     2581 gcagagatcc agtttggtta attaaggtac cgagggccta tttcccatga ttccttcata
     2641 tttgcatata cgatacaagg ctgttagaga gataattaga attaatttga ctgtaaacac
     2701 aaagatatta gtacaaaata cgtgacgtag aaagtaataa tttcttgggt agtttgcagt
     2761 tttaaaatta tgttttaaaa tggactatca tatgcttacc gtaacttgaa agtatttcga
     2821 tttcttggct ttatatatct tgtggaaagg acgaaacacc gtcaccgaga caccactgga
     2881 gttttagagc tagaaatagc aagttaaaat aaggctagtc cgttatcaac ttgaaaaagt
     2941 ggcaccgagt cggtgctttt ttgaattcgc tagctaggtc ttgaaaggag tgggaattgg
     3001 ctccggtgcc cgtcagtggg cagagcgcac atcgcccaca gtccccgaga agttgggggg
     3061 aggggtcggc aattgatccg gtgcctagag aaggtggcgc ggggtaaact gggaaagtga
     3121 tgtcgtgtac tggctccgcc tttttcccga gggtggggga gaaccgtata taagtgcagt
     3181 agtcgccgtg aacgttcttt ttcgcaacgg gtttgccgcc agaacacagg accggttcta
     3241 gagcgctgcc accatggaca agaagtacag catcggcctg gacatcggca ccaactctgt
     3301 gggctgggcc gtgatcaccg acgagtacaa ggtgcccagc aagaaattca aggtgctggg
     3361 caacaccgac cggcacagca tcaagaagaa cctgatcgga gccctgctgt tcgacagcgg
     3421 cgaaacagcc gaggccaccc ggctgaagag aaccgccaga agaagataca ccagacggaa
     3481 gaaccggatc tgctatctgc aagagatctt cagcaacgag atggccaagg tggacgacag
     3541 cttcttccac agactggaag agtccttcct ggtggaagag gataagaagc acgagcggca
     3601 ccccatcttc ggcaacatcg tggacgaggt ggcctaccac gagaagtacc ccaccatcta
     3661 ccacctgaga aagaaactgg tggacagcac cgacaaggcc gacctgcggc tgatctatct
     3721 ggccctggcc cacatgatca agttccgggg ccacttcctg atcgagggcg acctgaaccc
     3781 cgacaacagc gacgtggaca agctgttcat ccagctggtg cagacctaca accagctgtt
     3841 cgaggaaaac cccatcaacg ccagcggcgt ggacgccaag gccatcctgt ctgccagact
     3901 gagcaagagc agacggctgg aaaatctgat cgcccagctg cccggcgaga agaagaatgg
     3961 cctgttcgga aacctgattg ccctgagcct gggcctgacc cccaacttca agagcaactt
     4021 cgacctggcc gaggatgcca aactgcagct gagcaaggac acctacgacg acgacctgga
     4081 caacctgctg gcccagatcg gcgaccagta cgccgacctg tttctggccg ccaagaacct
     4141 gtccgacgcc atcctgctga gcgacatcct gagagtgaac accgagatca ccaaggcccc
     4201 cctgagcgcc tctatgatca agagatacga cgagcaccac caggacctga ccctgctgaa
     4261 agctctcgtg cggcagcagc tgcctgagaa gtacaaagag attttcttcg accagagcaa
     4321 gaacggctac gccggctaca ttgacggcgg agccagccag gaagagttct acaagttcat
     4381 caagcccatc ctggaaaaga tggacggcac cgaggaactg ctcgtgaagc tgaacagaga
     4441 ggacctgctg cggaagcagc ggaccttcga caacggcagc atcccccacc agatccacct
     4501 gggagagctg cacgccattc tgcggcggca ggaagatttt tacccattcc tgaaggacaa
     4561 ccgggaaaag atcgagaaga tcctgacctt ccgcatcccc tactacgtgg gccctctggc
     4621 caggggaaac agcagattcg cctggatgac cagaaagagc gaggaaacca tcaccccctg
     4681 gaacttcgag gaagtggtgg acaagggcgc ttccgcccag agcttcatcg agcggatgac
     4741 caacttcgat aagaacctgc ccaacgagaa ggtgctgccc aagcacagcc tgctgtacga
     4801 gtacttcacc gtgtataacg agctgaccaa agtgaaatac gtgaccgagg gaatgagaaa
     4861 gcccgccttc ctgagcggcg agcagaaaaa ggccatcgtg gacctgctgt tcaagaccaa
     4921 ccggaaagtg accgtgaagc agctgaaaga ggactacttc aagaaaatcg agtgcttcga
     4981 ctccgtggaa atctccggcg tggaagatcg gttcaacgcc tccctgggca cataccacga
     5041 tctgctgaaa attatcaagg acaaggactt cctggacaat gaggaaaacg aggacattct
     5101 ggaagatatc gtgctgaccc tgacactgtt tgaggacaga gagatgatcg aggaacggct
     5161 gaaaacctat gcccacctgt tcgacgacaa agtgatgaag cagctgaagc ggcggagata
     5221 caccggctgg ggcaggctga gccggaagct gatcaacggc atccgggaca agcagtccgg
     5281 caagacaatc ctggatttcc tgaagtccga cggcttcgcc aacagaaact tcatgcagct
     5341 gatccacgac gacagcctga cctttaaaga ggacatccag aaagcccagg tgtccggcca
     5401 gggcgatagc ctgcacgagc acattgccaa tctggccggc agccccgcca ttaagaaggg
     5461 catcctgcag acagtgaagg tggtggacga gctcgtgaaa gtgatgggcc ggcacaagcc
     5521 cgagaacatc gtgatcgaaa tggccagaga gaaccagacc acccagaagg gacagaagaa
     5581 cagccgcgag agaatgaagc ggatcgaaga gggcatcaaa gagctgggca gccagatcct
     5641 gaaagaacac cccgtggaaa acacccagct gcagaacgag aagctgtacc tgtactacct
     5701 gcagaatggg cgggatatgt acgtggacca ggaactggac atcaaccggc tgtccgacta
     5761 cgatgtggac catatcgtgc ctcagagctt tctgaaggac gactccatcg acaacaaggt
     5821 gctgaccaga agcgacaaga accggggcaa gagcgacaac gtgccctccg aagaggtcgt
     5881 gaagaagatg aagaactact ggcggcagct gctgaacgcc aagctgatta cccagagaaa
     5941 gttcgacaat ctgaccaagg ccgagagagg cggcctgagc gaactggata aggccggctt
     6001 catcaagaga cagctggtgg aaacccggca gatcacaaag cacgtggcac agatcctgga
     6061 ctcccggatg aacactaagt acgacgagaa tgacaagctg atccgggaag tgaaagtgat
     6121 caccctgaag tccaagctgg tgtccgattt ccggaaggat ttccagtttt acaaagtgcg
     6181 cgagatcaac aactaccacc acgcccacga cgcctacctg aacgccgtcg tgggaaccgc
     6241 cctgatcaaa aagtacccta agctggaaag cgagttcgtg tacggcgact acaaggtgta
     6301 cgacgtgcgg aagatgatcg ccaagagcga gcaggaaatc ggcaaggcta ccgccaagta
     6361 cttcttctac agcaacatca tgaacttttt caagaccgag attaccctgg ccaacggcga
     6421 gatccggaag cggcctctga tcgagacaaa cggcgaaacc ggggagatcg tgtgggataa
     6481 gggccgggat tttgccaccg tgcggaaagt gctgagcatg ccccaagtga atatcgtgaa
     6541 aaagaccgag gtgcagacag gcggcttcag caaagagtct atcctgccca agaggaacag
     6601 cgataagctg atcgccagaa agaaggactg ggaccctaag aagtacggcg gcttcgacag
     6661 ccccaccgtg gcctattctg tgctggtggt ggccaaagtg gaaaagggca agtccaagaa
     6721 actgaagagt gtgaaagagc tgctggggat caccatcatg gaaagaagca gcttcgagaa
     6781 gaatcccatc gactttctgg aagccaaggg ctacaaagaa gtgaaaaagg acctgatcat
     6841 caagctgcct aagtactccc tgttcgagct ggaaaacggc cggaagagaa tgctggcctc
     6901 tgccggcgaa ctgcagaagg gaaacgaact ggccctgccc tccaaatatg tgaacttcct
     6961 gtacctggcc agccactatg agaagctgaa gggctccccc gaggataatg agcagaaaca
     7021 gctgtttgtg gaacagcaca agcactacct ggacgagatc atcgagcaga tcagcgagtt
     7081 ctccaagaga gtgatcctgg ccgacgctaa tctggacaaa gtgctgtccg cctacaacaa
     7141 gcaccgggat aagcccatca gagagcaggc cgagaatatc atccacctgt ttaccctgac
     7201 caatctggga gcccctgccg ccttcaagta ctttgacacc accatcgacc ggaagaggta
     7261 caccagcacc aaagaggtgc tggacgccac cctgatccac cagagcatca ccggcctgta
     7321 cgagacacgg atcgacctgt ctcagctggg aggcgacaag cgacctgccg ccacaaagaa
     7381 ggctggacag gctaagaaga agaaagatta caaagacgat gacgataagg gatccggcgc
     7441 aacaaacttc tctctgctga aacaagccgg agatgtcgaa gagaatcctg gaccgaaaaa
     7501 gcctgaactc accgcgacgt ctgtcgagaa gtttctgatc gaaaagttcg acagcgtgtc
     7561 cgacctgatg cagctctcgg agggcgaaga atctcgtgct ttcagcttcg atgtaggagg
     7621 gcgtggatat gtcctgcggg taaatagctg cgccgatggt ttctacaaag atcgttatgt
     7681 ttatcggcac tttgcatcgg ccgcgctccc gattccggaa gtgcttgaca ttggggaatt
     7741 cagcgagagc ctgacctatt gcatctcccg ccgtgcacag ggtgtcacgt tgcaagacct
     7801 gcctgaaacc gaactgcccg ctgttctgca gccggtcgcg gaggccatgg atgcgatcgc
     7861 tgcggccgat cttagccaga cgagcgggtt cggcccattc ggaccgcaag gaatcggtca
     7921 atacactaca tggcgtgatt tcatatgcgc gattgctgat ccccatgtgt atcactggca
     7981 aactgtgatg gacgacaccg tcagtgcgtc cgtcgcgcag gctctcgatg agctgatgct
     8041 ttgggccgag gactgccccg aagtccggca cctcgtgcac gcggatttcg gctccaacaa
     8101 tgtcctgacg gacaatggcc gcataacagc ggtcattgac tggagcgagg cgatgttcgg
     8161 ggattcccaa tacgaggtcg ccaacatctt cttctggagg ccgtggttgg cttgtatgga
     8221 gcagcagacg cgctacttcg agcggaggca tccggagctt gcaggatcgc cgcggctccg
     8281 ggcgtatatg ctccgcattg gtcttgacca actctatcag agcttggttg acggcaattt
     8341 cgatgatgca gcttgggcgc agggtcgatg cgacgcaatc gtccgatccg gagccgggac
     8401 tgtcgggcgt acacaaatcg cccgcagaag cgcggccgtc tggaccgatg gctgtgtaga
     8461 agtacttcgc cgatagacgc gttaagtcga caatcaacct ctggattaca aaatttgtga
     8521 aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt
     8581 aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa
     8641 atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt
     8701 gtgcactgtg tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct
     8761 cctttccggg actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg
     8821 ccttgcccgc tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc
     8881 ggggaaatca tcgtcctttc cttggctgct cgcctgtgtt gccacctgga ttctgcgcgg
     8941 gacgtccttc tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct
     9001 gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga gtcggatctc
     9061 cctttgggcc gcctccccgc gtcgacttta agaccaatga cttacaaggc agctgtagat
     9121 cttagccact ttttaaaaga aaagggggga ctggaagggc taattcactc ccaacgaaga
     9181 caagatctgc tttttgcttg tactgggtct ctctggttag accagatctg agcctgggag
     9241 ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc ttgagtgctt
     9301 caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct cagacccttt
     9361 tagtcagtgt ggaaaatctc tagcagggcc cgtttaaacc cgctgatcag cctcgactgt
     9421 gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct tgaccctgga
     9481 aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc attgtctgag
     9541 taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg aggattggga
     9601 agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg cggaaagaac
     9661 cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa gcgcggcggg
     9721 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     9781 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     9841 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     9901 ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     9961 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
    10021 tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
    10081 aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt gtgtcagtta
    10141 gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat gcatctcaat
    10201 tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
    10261 atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta
    10321 actccgccca gttccgccca ttctccgccc catggctgac taattttttt tatttatgca
    10381 gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga
    10441 ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg atctgatcag
    10501 cacgtgttga caattaatca tcggcatagt atatcggcat agtataatac gacaaggtga
    10561 ggaactaaac catggccaag ttgaccagtg ccgttccggt gctcaccgcg cgcgacgtcg
    10621 ccggagcggt cgagttctgg accgaccggc tcgggttctc ccgggacttc gtggaggacg
    10681 acttcgccgg tgtggtccgg gacgacgtga ccctgttcat cagcgcggtc caggaccagg
    10741 tggtgccgga caacaccctg gcctgggtgt gggtgcgcgg cctggacgag ctgtacgccg
    10801 agtggtcgga ggtcgtgtcc acgaacttcc gggacgcctc cgggccggcc atgaccgaga
    10861 tcggcgagca gccgtggggg cgggagttcg ccctgcgcga cccggccggc aactgcgtgc
    10921 acttcgtggc cgaggagcag gactgacacg tgctacgaga tttcgattcc accgccgcct
    10981 tctatgaaag gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc
    11041 gcggggatct catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg
    11101 gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt
    11161 ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct
    11221 ctagctagag cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
    11281 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
    11341 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
    11401 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
    11461 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
    11521 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
    11581 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
    11641 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
    11701 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
    11761 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
    11821 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
    11881 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
    11941 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
    12001 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
    12061 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
    12121 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
    12181 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
    12241 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
    12301 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
    12361 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
    12421 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
    12481 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
    12541 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
    12601 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
    12661 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
    12721 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
    12781 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
    12841 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
    12901 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
    12961 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
    13021 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
    13081 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
    13141 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
    13201 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
    13261 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
    13321 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
    13381 cccgaaaagt gccacctgac
//
