LOCUS       sequence-431999-       13023 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Knockout vector for mouse Plekha8 (Fapp2).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13023)
  AUTHORS   Hollingsworth LR, Veeraraghavan P, Paulo JA, Harper JW
  TITLE     Spatiotemporal proteomic profiling of cellular responses to NLRP3 
            agonists
  JOURNAL   bioRxiv 2024.04.19.590338
REFERENCE   2  (bases 1 to 13023)
  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: "bioRxiv 
            2024.04.19.590338"
COMMENT     Sequence Label: msPlekha8 (msFapp2) g1 lentiCRISPRv2-opti
FEATURES             Location/Qualifiers
     source          1..13023
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        382..580
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             598..778
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    825..950
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1443..1676
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1861..1905
                     /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    2203..2320
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2371..2619
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2371..2391
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2542..2561
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     promoter        2788..2999
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      3018..3027
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3024..7127
                     /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             3024..7127
                     /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             7128..7175
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7176..7199
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             7209..7265
                     /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             7266..7862
                     /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     7759..7779
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    7878..8466
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7931..7951)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8349..8360)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             8538..8771
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(8797..8814)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    8803..9027
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      9073..9501
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9160..9179)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9370..9391
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(9510..9530)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        9515..9844
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      9695..9830
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     9757..9776
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        9892..9939
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             9958..10332
                     /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    10462..10583
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(10499..10518)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     10553..10572
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(10632..10648)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(10632..10648)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(10645..10667)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    10656..10672
                     /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(10680..10686,10687..10704,10705..10710))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    10725..10746
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(10863..10880)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11034..11622)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11114..11133)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(11793..12584,12585..12653))
                     /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     12416..12435
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(12654..12758)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(12833..12852)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta gttcatagcc
       61 catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc tgaccgccca
      121 acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg ccaataggga
      181 ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg gcagtacatc
      241 aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct
      301 ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac atctacgtat
      361 tagtcatcgc tattaccatg gtgatgcggt tttggcagta catcaatggg cgtggatagc
      421 ggtttgactc acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt
      481 ggcaccaaaa tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa
      541 tgggcggtag gcgtgtacgg tgggaggtct atataagcag cgcgttttgc ctgtactggg
      601 tctctctggt tagaccagat ctgagcctgg gagctctctg gctaactagg gaacccactg
      661 cttaagcctc aataaagctt gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt
      721 gactctggta actagagatc cctcagaccc ttttagtcag tgtggaaaat ctctagcagt
      781 ggcgcccgaa cagggacttg aaagcgaaag ggaaaccaga ggagctctct cgacgcagga
      841 ctcggcttgc tgaagcgcgc acggcaagag gcgaggggcg gcgactggtg agtacgccaa
      901 aaattttgac tagcggaggc tagaaggaga gagatgggtg cgagagcgtc agtattaagc
      961 gggggagaat tagatcgcga tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat
     1021 ataaattaaa acatatagta tgggcaagca gggagctaga acgattcgca gttaatcctg
     1081 gcctgttaga aacatcagaa ggctgtagac aaatactggg acagctacaa ccatcccttc
     1141 agacaggatc agaagaactt agatcattat ataatacagt agcaaccctc tattgtgtgc
     1201 atcaaaggat agagataaaa gacaccaagg aagctttaga caagatagag gaagagcaaa
     1261 acaaaagtaa gaccaccgca cagcaagcgg ccgctgatct tcagacctgg aggaggagat
     1321 atgagggaca attggagaag tgaattatat aaatataaag tagtaaaaat tgaaccatta
     1381 ggagtagcac ccaccaaggc aaagagaaga gtggtgcaga gagaaaaaag agcagtggga
     1441 ataggagctt tgttccttgg gttcttggga gcagcaggaa gcactatggg cgcagcgtca
     1501 atgacgctga cggtacaggc cagacaatta ttgtctggta tagtgcagca gcagaacaat
     1561 ttgctgaggg ctattgaggc gcaacagcat ctgttgcaac tcacagtctg gggcatcaag
     1621 cagctccagg caagaatcct ggctgtggaa agatacctaa aggatcaaca gctcctgggg
     1681 atttggggtt gctctggaaa actcatttgc accactgctg tgccttggaa tgctagttgg
     1741 agtaataaat ctctggaaca gatttggaat cacacgacct ggatggagtg ggacagagaa
     1801 attaacaatt acacaagctt aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa
     1861 aagaatgaac aagaattatt ggaattagat aaatgggcaa gtttgtggaa ttggtttaac
     1921 ataacaaatt ggctgtggta tataaaatta ttcataatga tagtaggagg cttggtaggt
     1981 ttaagaatag tttttgctgt actttctata gtgaatagag ttaggcaggg atattcacca
     2041 ttatcgtttc agacccacct cccaaccccg aggggacccg acaggcccga aggaatagaa
     2101 gaagaaggtg gagagagaga cagagacaga tccattcgat tagtgaacgg atcggcactg
     2161 cgtgcgccaa ttctgcagac aaatggcagt attcatccac aattttaaaa gaaaaggggg
     2221 gattgggggg tacagtgcag gggaaagaat agtagacata atagcaacag acatacaaac
     2281 taaagaatta caaaaacaaa ttacaaaaat tcaaaatttt cgggtttatt acagggacag
     2341 cagagatcca gtttggttaa ttaaggtacc gagggcctat ttcccatgat tccttcatat
     2401 ttgcatatac gatacaaggc tgttagagag ataattagaa ttaatttgac tgtaaacaca
     2461 aagatattag tacaaaatac gtgacgtaga aagtaataat ttcttgggta gtttgcagtt
     2521 ttaaaattat gttttaaaat ggactatcat atgcttaccg taacttgaaa gtatttcgat
     2581 ttcttggctt tatatatctt gtggaaagga cgaaacaccg cgtcgctcag gttaggaact
     2641 gtttaagagc tatgctggaa acagcatagc aagtttaaat aaggctagtc cgttatcaac
     2701 ttgaaaaagt ggcaccgagt cggtgctttt ttgaattcgc tagctaggtc ttgaaaggag
     2761 tgggaattgg ctccggtgcc cgtcagtggg cagagcgcac atcgcccaca gtccccgaga
     2821 agttgggggg aggggtcggc aattgatccg gtgcctagag aaggtggcgc ggggtaaact
     2881 gggaaagtga tgtcgtgtac tggctccgcc tttttcccga gggtggggga gaaccgtata
     2941 taagtgcagt agtcgccgtg aacgttcttt ttcgcaacgg gtttgccgcc agaacacagg
     3001 accggttcta gagcgctgcc accatggaca agaagtacag catcggcctg gacatcggca
     3061 ccaactctgt gggctgggcc gtgatcaccg acgagtacaa ggtgcccagc aagaaattca
     3121 aggtgctggg caacaccgac cggcacagca tcaagaagaa cctgatcgga gccctgctgt
     3181 tcgacagcgg cgaaacagcc gaggccaccc ggctgaagag aaccgccaga agaagataca
     3241 ccagacggaa gaaccggatc tgctatctgc aagagatctt cagcaacgag atggccaagg
     3301 tggacgacag cttcttccac agactggaag agtccttcct ggtggaagag gataagaagc
     3361 acgagcggca ccccatcttc ggcaacatcg tggacgaggt ggcctaccac gagaagtacc
     3421 ccaccatcta ccacctgaga aagaaactgg tggacagcac cgacaaggcc gacctgcggc
     3481 tgatctatct ggccctggcc cacatgatca agttccgggg ccacttcctg atcgagggcg
     3541 acctgaaccc cgacaacagc gacgtggaca agctgttcat ccagctggtg cagacctaca
     3601 accagctgtt cgaggaaaac cccatcaacg ccagcggcgt ggacgccaag gccatcctgt
     3661 ctgccagact gagcaagagc agacggctgg aaaatctgat cgcccagctg cccggcgaga
     3721 agaagaatgg cctgttcgga aacctgattg ccctgagcct gggcctgacc cccaacttca
     3781 agagcaactt cgacctggcc gaggatgcca aactgcagct gagcaaggac acctacgacg
     3841 acgacctgga caacctgctg gcccagatcg gcgaccagta cgccgacctg tttctggccg
     3901 ccaagaacct gtccgacgcc atcctgctga gcgacatcct gagagtgaac accgagatca
     3961 ccaaggcccc cctgagcgcc tctatgatca agagatacga cgagcaccac caggacctga
     4021 ccctgctgaa agctctcgtg cggcagcagc tgcctgagaa gtacaaagag attttcttcg
     4081 accagagcaa gaacggctac gccggctaca ttgacggcgg agccagccag gaagagttct
     4141 acaagttcat caagcccatc ctggaaaaga tggacggcac cgaggaactg ctcgtgaagc
     4201 tgaacagaga ggacctgctg cggaagcagc ggaccttcga caacggcagc atcccccacc
     4261 agatccacct gggagagctg cacgccattc tgcggcggca ggaagatttt tacccattcc
     4321 tgaaggacaa ccgggaaaag atcgagaaga tcctgacctt ccgcatcccc tactacgtgg
     4381 gccctctggc caggggaaac agcagattcg cctggatgac cagaaagagc gaggaaacca
     4441 tcaccccctg gaacttcgag gaagtggtgg acaagggcgc ttccgcccag agcttcatcg
     4501 agcggatgac caacttcgat aagaacctgc ccaacgagaa ggtgctgccc aagcacagcc
     4561 tgctgtacga gtacttcacc gtgtataacg agctgaccaa agtgaaatac gtgaccgagg
     4621 gaatgagaaa gcccgccttc ctgagcggcg agcagaaaaa ggccatcgtg gacctgctgt
     4681 tcaagaccaa ccggaaagtg accgtgaagc agctgaaaga ggactacttc aagaaaatcg
     4741 agtgcttcga ctccgtggaa atctccggcg tggaagatcg gttcaacgcc tccctgggca
     4801 cataccacga tctgctgaaa attatcaagg acaaggactt cctggacaat gaggaaaacg
     4861 aggacattct ggaagatatc gtgctgaccc tgacactgtt tgaggacaga gagatgatcg
     4921 aggaacggct gaaaacctat gcccacctgt tcgacgacaa agtgatgaag cagctgaagc
     4981 ggcggagata caccggctgg ggcaggctga gccggaagct gatcaacggc atccgggaca
     5041 agcagtccgg caagacaatc ctggatttcc tgaagtccga cggcttcgcc aacagaaact
     5101 tcatgcagct gatccacgac gacagcctga cctttaaaga ggacatccag aaagcccagg
     5161 tgtccggcca gggcgatagc ctgcacgagc acattgccaa tctggccggc agccccgcca
     5221 ttaagaaggg catcctgcag acagtgaagg tggtggacga gctcgtgaaa gtgatgggcc
     5281 ggcacaagcc cgagaacatc gtgatcgaaa tggccagaga gaaccagacc acccagaagg
     5341 gacagaagaa cagccgcgag agaatgaagc ggatcgaaga gggcatcaaa gagctgggca
     5401 gccagatcct gaaagaacac cccgtggaaa acacccagct gcagaacgag aagctgtacc
     5461 tgtactacct gcagaatggg cgggatatgt acgtggacca ggaactggac atcaaccggc
     5521 tgtccgacta cgatgtggac catatcgtgc ctcagagctt tctgaaggac gactccatcg
     5581 acaacaaggt gctgaccaga agcgacaaga accggggcaa gagcgacaac gtgccctccg
     5641 aagaggtcgt gaagaagatg aagaactact ggcggcagct gctgaacgcc aagctgatta
     5701 cccagagaaa gttcgacaat ctgaccaagg ccgagagagg cggcctgagc gaactggata
     5761 aggccggctt catcaagaga cagctggtgg aaacccggca gatcacaaag cacgtggcac
     5821 agatcctgga ctcccggatg aacactaagt acgacgagaa tgacaagctg atccgggaag
     5881 tgaaagtgat caccctgaag tccaagctgg tgtccgattt ccggaaggat ttccagtttt
     5941 acaaagtgcg cgagatcaac aactaccacc acgcccacga cgcctacctg aacgccgtcg
     6001 tgggaaccgc cctgatcaaa aagtacccta agctggaaag cgagttcgtg tacggcgact
     6061 acaaggtgta cgacgtgcgg aagatgatcg ccaagagcga gcaggaaatc ggcaaggcta
     6121 ccgccaagta cttcttctac agcaacatca tgaacttttt caagaccgag attaccctgg
     6181 ccaacggcga gatccggaag cggcctctga tcgagacaaa cggcgaaacc ggggagatcg
     6241 tgtgggataa gggccgggat tttgccaccg tgcggaaagt gctgagcatg ccccaagtga
     6301 atatcgtgaa aaagaccgag gtgcagacag gcggcttcag caaagagtct atcctgccca
     6361 agaggaacag cgataagctg atcgccagaa agaaggactg ggaccctaag aagtacggcg
     6421 gcttcgacag ccccaccgtg gcctattctg tgctggtggt ggccaaagtg gaaaagggca
     6481 agtccaagaa actgaagagt gtgaaagagc tgctggggat caccatcatg gaaagaagca
     6541 gcttcgagaa gaatcccatc gactttctgg aagccaaggg ctacaaagaa gtgaaaaagg
     6601 acctgatcat caagctgcct aagtactccc tgttcgagct ggaaaacggc cggaagagaa
     6661 tgctggcctc tgccggcgaa ctgcagaagg gaaacgaact ggccctgccc tccaaatatg
     6721 tgaacttcct gtacctggcc agccactatg agaagctgaa gggctccccc gaggataatg
     6781 agcagaaaca gctgtttgtg gaacagcaca agcactacct ggacgagatc atcgagcaga
     6841 tcagcgagtt ctccaagaga gtgatcctgg ccgacgctaa tctggacaaa gtgctgtccg
     6901 cctacaacaa gcaccgggat aagcccatca gagagcaggc cgagaatatc atccacctgt
     6961 ttaccctgac caatctggga gcccctgccg ccttcaagta ctttgacacc accatcgacc
     7021 ggaagaggta caccagcacc aaagaggtgc tggacgccac cctgatccac cagagcatca
     7081 ccggcctgta cgagacacgg atcgacctgt ctcagctggg aggcgacaag cgacctgccg
     7141 ccacaaagaa ggctggacag gctaagaaga agaaagatta caaagacgat gacgataagg
     7201 gatccggcgc aacaaacttc tctctgctga aacaagccgg agatgtcgaa gagaatcctg
     7261 gaccgaccga gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc cccagggccg
     7321 tacgcaccct cgccgccgcg ttcgccgact accccgccac gcgccacacc gtcgatccgg
     7381 accgccacat cgagcgggtc accgagctgc aagaactctt cctcacgcgc gtcgggctcg
     7441 acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg accacgccgg
     7501 agagcgtcga agcgggggcg gtgttcgccg agatcggccc gcgcatggcc gagttgagcg
     7561 gttcccggct ggccgcgcag caacagatgg aaggcctcct ggcgccgcac cggcccaagg
     7621 agcccgcgtg gttcctggcc accgtcggag tctcgcccga ccaccagggc aagggtctgg
     7681 gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg cgccggggtg cccgccttcc
     7741 tggagacctc cgcgccccgc aacctcccct tctacgagcg gctcggcttc accgtcaccg
     7801 ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag cccggtgcct
     7861 gaacgcgtta agtcgacaat caacctctgg attacaaaat ttgtgaaaga ttgactggta
     7921 ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc
     7981 atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt
     8041 ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg
     8101 ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt
     8161 tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct
     8221 ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg aaatcatcgt
     8281 cctttccttg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg tccttctgct
     8341 acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc
     8401 ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg gatctccctt tgggccgcct
     8461 ccccgcgtcg actttaagac caatgactta caaggcagct gtagatctta gccacttttt
     8521 aaaagaaaag gggggactgg aagggctaat tcactcccaa cgaagacaag atctgctttt
     8581 tgcttgtact gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact
     8641 agggaaccca ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc
     8701 ccgtctgttg tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa
     8761 aatctctagc agggcccgtt taaacccgct gatcagcctc gactgtgcct tctagttgcc
     8821 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
     8881 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
     8941 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc
     9001 atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta
     9061 gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     9121 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     9181 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     9241 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     9301 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     9361 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     9421 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     9481 aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc
     9541 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag
     9601 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     9661 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
     9721 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     9781 ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
     9841 caaaaagctc ccgggagctt gtatatccat tttcggatct gatcagcacg tgttgacaat
     9901 taatcatcgg catagtatat cggcatagta taatacgaca aggtgaggaa ctaaaccatg
     9961 gccaagttga ccagtgccgt tccggtgctc accgcgcgcg acgtcgccgg agcggtcgag
    10021 ttctggaccg accggctcgg gttctcccgg gacttcgtgg aggacgactt cgccggtgtg
    10081 gtccgggacg acgtgaccct gttcatcagc gcggtccagg accaggtggt gccggacaac
    10141 accctggcct gggtgtgggt gcgcggcctg gacgagctgt acgccgagtg gtcggaggtc
    10201 gtgtccacga acttccggga cgcctccggg ccggccatga ccgagatcgg cgagcagccg
    10261 tgggggcggg agttcgccct gcgcgacccg gccggcaact gcgtgcactt cgtggccgag
    10321 gagcaggact gacacgtgct acgagatttc gattccaccg ccgccttcta tgaaaggttg
    10381 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
    10441 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
    10501 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
    10561 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
    10621 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
    10681 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
    10741 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
    10801 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
    10861 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
    10921 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
    10981 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
    11041 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
    11101 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
    11161 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
    11221 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
    11281 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
    11341 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
    11401 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
    11461 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
    11521 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
    11581 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
    11641 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
    11701 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
    11761 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
    11821 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
    11881 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
    11941 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
    12001 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
    12061 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
    12121 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
    12181 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
    12241 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
    12301 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
    12361 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
    12421 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
    12481 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
    12541 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
    12601 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
    12661 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
    12721 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
    12781 cctgacgtcg acggatcggg agatctcccg atcccctatg gtgcactctc agtacaatct
    12841 gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg
    12901 agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac aattgcatga
    12961 agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc
    13021 gtt
//
