LOCUS       sequence-465545-       13212 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Knock out of murine ITGA5.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13212)
  AUTHORS   Nikolatou K, Sandilands E, Roman-Fernandez A, Cumming EM, Freckmann 
            E, Lilla S, Buetow L, McGarry L, Neilson M, Shaw R, Strachan D, 
            Miller C, Huang DT, McNeish IA, Norman JC, Zanivan S, Bryant DM
  TITLE     PTEN deficiency exposes a requirement for an ARF GTPase module for 
            integrin-dependent invasion in ovarian cancer.
  JOURNAL   EMBO J. 2023 Sep 18;42(18):e113987. doi: 10.15252/embj.2023113987. 
            Epub 2023 Aug 14.
  PUBMED    37577760
REFERENCE   2  (bases 1 to 13212)
  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: "EMBO J."; 
            date: "2023-09-18"; volume: "42(18):e113987. doi"; pages: " 
            10.15252/embj.2023113987. Epub 2023 Aug 14"
COMMENT     Sequence Label: pLentiCRISPR V2 Neo sgITGA5_1
FEATURES             Location/Qualifiers
     source          1..13212
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     532..551
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     misc_feature    678..1266
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(731..751)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1149..1160)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             1338..1571
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(1597..1614)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1603..1827
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1873..2301
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1960..1979)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2170..2191
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2310..2330)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2315..2644
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2495..2630
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2557..2576
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        2692..2739
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             2758..3132
                     /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    3262..3383
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3299..3318)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3353..3372
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3432..3448)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3432..3448)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3445..3467)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3456..3472
                     /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(3480..3486,3487..3504,3505..3510))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3525..3546
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3663..3680)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3834..4422)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3914..3933)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4593..5384,5385..5453))
                     /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     5216..5235
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5454..5558)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5633..5652)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        5828..6207
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        6209..6407
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     6362..6382
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             6425..6605
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    6652..6777
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    7270..7503
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             7688..7732
                     /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    8030..8147
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        8198..8446
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     8198..8218
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     8369..8388
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        8468..8543
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        8605..8816
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      8835..8844
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             8841..12944
                     /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             8841..12944
                     /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             12945..12992
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             12993..13016
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             13026..13082
                     /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             join(13083..13212,1..662)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="IEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRPV
                     LFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLSS
                     HLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQG
                     LAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIAL
                     ATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(13134..13153)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
ORIGIN
        1 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcag gacgaggcag
       61 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
      121 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
      181 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
      241 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
      301 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
      361 tcgcgccagc cgaactgttc gccaggctca aggcgcgcat gcccgacggc gaggatctcg
      421 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
      481 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
      541 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
      601 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
      661 gaacgcgtta agtcgacaat caacctctgg attacaaaat ttgtgaaaga ttgactggta
      721 ttcttaacta tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc
      781 atgctattgc ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt
      841 ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg
      901 ctgacgcaac ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt
      961 tcgctttccc cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct
     1021 ggacaggggc tcggctgttg ggcactgaca attccgtggt gttgtcgggg aaatcatcgt
     1081 cctttccttg gctgctcgcc tgtgttgcca cctggattct gcgcgggacg tccttctgct
     1141 acgtcccttc ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc
     1201 ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg gatctccctt tgggccgcct
     1261 ccccgcgtcg actttaagac caatgactta caaggcagct gtagatctta gccacttttt
     1321 aaaagaaaag gggggactgg aagggctaat tcactcccaa cgaagacaag atctgctttt
     1381 tgcttgtact gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact
     1441 agggaaccca ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc
     1501 ccgtctgttg tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa
     1561 aatctctagc agggcccgtt taaacccgct gatcagcctc gactgtgcct tctagttgcc
     1621 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
     1681 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
     1741 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc
     1801 atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta
     1861 gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     1921 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     1981 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     2041 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     2101 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     2161 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     2221 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     2281 aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc
     2341 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag
     2401 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     2461 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
     2521 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     2581 ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
     2641 caaaaagctc ccgggagctt gtatatccat tttcggatct gatcagcacg tgttgacaat
     2701 taatcatcgg catagtatat cggcatagta taatacgaca aggtgaggaa ctaaaccatg
     2761 gccaagttga ccagtgccgt tccggtgctc accgcgcgcg acgtcgccgg agcggtcgag
     2821 ttctggaccg accggctcgg gttctcccgg gacttcgtgg aggacgactt cgccggtgtg
     2881 gtccgggacg acgtgaccct gttcatcagc gcggtccagg accaggtggt gccggacaac
     2941 accctggcct gggtgtgggt gcgcggcctg gacgagctgt acgccgagtg gtcggaggtc
     3001 gtgtccacga acttccggga cgcctccggg ccggccatga ccgagatcgg cgagcagccg
     3061 tgggggcggg agttcgccct gcgcgacccg gccggcaact gcgtgcactt cgtggccgag
     3121 gagcaggact gacacgtgct acgagatttc gattccaccg ccgccttcta tgaaaggttg
     3181 ggcttcggaa tcgttttccg ggacgccggc tggatgatcc tccagcgcgg ggatctcatg
     3241 ctggagttct tcgcccaccc caacttgttt attgcagctt ataatggtta caaataaagc
     3301 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     3361 tccaaactca tcaatgtatc ttatcatgtc tgtataccgt cgacctctag ctagagcttg
     3421 gcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt atccgctcac aattccacac
     3481 aacatacgag ccggaagcat aaagtgtaaa gcctggggtg cctaatgagt gagctaactc
     3541 acattaattg cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg
     3601 cattaatgaa tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ctcttccgct
     3661 tcctcgctca ctgactcgct gcgctcggtc gttcggctgc ggcgagcggt atcagctcac
     3721 tcaaaggcgg taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga
     3781 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     3841 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     3901 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     3961 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     4021 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     4081 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     4141 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     4201 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     4261 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     4321 aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     4381 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     4441 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     4501 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     4561 taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct
     4621 atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata
     4681 actacgatac gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca
     4741 cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga
     4801 agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga
     4861 gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg
     4921 gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga
     4981 gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt
     5041 gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct
     5101 cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca
     5161 ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat
     5221 accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga
     5281 aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc
     5341 aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg
     5401 caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc
     5461 ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt
     5521 gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca
     5581 cctgacgtcg acggatcggg agatctcccg atcccctatg gtgcactctc agtacaatct
     5641 gctctgatgc cgcatagtta agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg
     5701 agtagtgcgc gagcaaaatt taagctacaa caaggcaagg cttgaccgac aattgcatga
     5761 agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc
     5821 gcgcgttgac attgattatt gactagttat taatagtaat caattacggg gtcattagtt
     5881 catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga
     5941 ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca
     6001 atagggactt tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca
     6061 gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg
     6121 cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc
     6181 tacgtattag tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt
     6241 ggatagcggt ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt
     6301 ttgttttggc accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg
     6361 acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg
     6421 tactgggtct ctctggttag accagatctg agcctgggag ctctctggct aactagggaa
     6481 cccactgctt aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct
     6541 gttgtgtgac tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc
     6601 tagcagtggc gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga
     6661 cgcaggactc ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt
     6721 acgccaaaaa ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt
     6781 attaagcggg ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag
     6841 aaaaaatata aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt
     6901 aatcctggcc tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca
     6961 tcccttcaga caggatcaga agaacttaga tcattatata atacagtagc aaccctctat
     7021 tgtgtgcatc aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa
     7081 gagcaaaaca aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg
     7141 aggagatatg agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga
     7201 accattagga gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc
     7261 agtgggaata ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc
     7321 agcgtcaatg acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca
     7381 gaacaatttg ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg
     7441 catcaagcag ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct
     7501 cctggggatt tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc
     7561 tagttggagt aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga
     7621 cagagaaatt aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca
     7681 gcaagaaaag aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg
     7741 gtttaacata acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt
     7801 ggtaggttta agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata
     7861 ttcaccatta tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg
     7921 aatagaagaa gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc
     7981 ggcactgcgt gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa
     8041 aaggggggat tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca
     8101 tacaaactaa agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca
     8161 gggacagcag agatccagtt tggttaatta aggtaccgag ggcctatttc ccatgattcc
     8221 ttcatatttg catatacgat acaaggctgt tagagagata attagaatta atttgactgt
     8281 aaacacaaag atattagtac aaaatacgtg acgtagaaag taataatttc ttgggtagtt
     8341 tgcagtttta aaattatgtt ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta
     8401 tttcgatttc ttggctttat atatcttgtg gaaaggacga aacaccggtc ttacatagcc
     8461 ataggtggtt ttagagctag aaatagcaag ttaaaataag gctagtccgt tatcaacttg
     8521 aaaaagtggc accgagtcgg tgcttttttg aattcgctag ctaggtcttg aaaggagtgg
     8581 gaattggctc cggtgcccgt cagtgggcag agcgcacatc gcccacagtc cccgagaagt
     8641 tggggggagg ggtcggcaat tgatccggtg cctagagaag gtggcgcggg gtaaactggg
     8701 aaagtgatgt cgtgtactgg ctccgccttt ttcccgaggg tgggggagaa ccgtatataa
     8761 gtgcagtagt cgccgtgaac gttctttttc gcaacgggtt tgccgccaga acacaggacc
     8821 ggttctagag cgctgccacc atggacaaga agtacagcat cggcctggac atcggcacca
     8881 actctgtggg ctgggccgtg atcaccgacg agtacaaggt gcccagcaag aaattcaagg
     8941 tgctgggcaa caccgaccgg cacagcatca agaagaacct gatcggagcc ctgctgttcg
     9001 acagcggcga aacagccgag gccacccggc tgaagagaac cgccagaaga agatacacca
     9061 gacggaagaa ccggatctgc tatctgcaag agatcttcag caacgagatg gccaaggtgg
     9121 acgacagctt cttccacaga ctggaagagt ccttcctggt ggaagaggat aagaagcacg
     9181 agcggcaccc catcttcggc aacatcgtgg acgaggtggc ctaccacgag aagtacccca
     9241 ccatctacca cctgagaaag aaactggtgg acagcaccga caaggccgac ctgcggctga
     9301 tctatctggc cctggcccac atgatcaagt tccggggcca cttcctgatc gagggcgacc
     9361 tgaaccccga caacagcgac gtggacaagc tgttcatcca gctggtgcag acctacaacc
     9421 agctgttcga ggaaaacccc atcaacgcca gcggcgtgga cgccaaggcc atcctgtctg
     9481 ccagactgag caagagcaga cggctggaaa atctgatcgc ccagctgccc ggcgagaaga
     9541 agaatggcct gttcggaaac ctgattgccc tgagcctggg cctgaccccc aacttcaaga
     9601 gcaacttcga cctggccgag gatgccaaac tgcagctgag caaggacacc tacgacgacg
     9661 acctggacaa cctgctggcc cagatcggcg accagtacgc cgacctgttt ctggccgcca
     9721 agaacctgtc cgacgccatc ctgctgagcg acatcctgag agtgaacacc gagatcacca
     9781 aggcccccct gagcgcctct atgatcaaga gatacgacga gcaccaccag gacctgaccc
     9841 tgctgaaagc tctcgtgcgg cagcagctgc ctgagaagta caaagagatt ttcttcgacc
     9901 agagcaagaa cggctacgcc ggctacattg acggcggagc cagccaggaa gagttctaca
     9961 agttcatcaa gcccatcctg gaaaagatgg acggcaccga ggaactgctc gtgaagctga
    10021 acagagagga cctgctgcgg aagcagcgga ccttcgacaa cggcagcatc ccccaccaga
    10081 tccacctggg agagctgcac gccattctgc ggcggcagga agatttttac ccattcctga
    10141 aggacaaccg ggaaaagatc gagaagatcc tgaccttccg catcccctac tacgtgggcc
    10201 ctctggccag gggaaacagc agattcgcct ggatgaccag aaagagcgag gaaaccatca
    10261 ccccctggaa cttcgaggaa gtggtggaca agggcgcttc cgcccagagc ttcatcgagc
    10321 ggatgaccaa cttcgataag aacctgccca acgagaaggt gctgcccaag cacagcctgc
    10381 tgtacgagta cttcaccgtg tataacgagc tgaccaaagt gaaatacgtg accgagggaa
    10441 tgagaaagcc cgccttcctg agcggcgagc agaaaaaggc catcgtggac ctgctgttca
    10501 agaccaaccg gaaagtgacc gtgaagcagc tgaaagagga ctacttcaag aaaatcgagt
    10561 gcttcgactc cgtggaaatc tccggcgtgg aagatcggtt caacgcctcc ctgggcacat
    10621 accacgatct gctgaaaatt atcaaggaca aggacttcct ggacaatgag gaaaacgagg
    10681 acattctgga agatatcgtg ctgaccctga cactgtttga ggacagagag atgatcgagg
    10741 aacggctgaa aacctatgcc cacctgttcg acgacaaagt gatgaagcag ctgaagcggc
    10801 ggagatacac cggctggggc aggctgagcc ggaagctgat caacggcatc cgggacaagc
    10861 agtccggcaa gacaatcctg gatttcctga agtccgacgg cttcgccaac agaaacttca
    10921 tgcagctgat ccacgacgac agcctgacct ttaaagagga catccagaaa gcccaggtgt
    10981 ccggccaggg cgatagcctg cacgagcaca ttgccaatct ggccggcagc cccgccatta
    11041 agaagggcat cctgcagaca gtgaaggtgg tggacgagct cgtgaaagtg atgggccggc
    11101 acaagcccga gaacatcgtg atcgaaatgg ccagagagaa ccagaccacc cagaagggac
    11161 agaagaacag ccgcgagaga atgaagcgga tcgaagaggg catcaaagag ctgggcagcc
    11221 agatcctgaa agaacacccc gtggaaaaca cccagctgca gaacgagaag ctgtacctgt
    11281 actacctgca gaatgggcgg gatatgtacg tggaccagga actggacatc aaccggctgt
    11341 ccgactacga tgtggaccat atcgtgcctc agagctttct gaaggacgac tccatcgaca
    11401 acaaggtgct gaccagaagc gacaagaacc ggggcaagag cgacaacgtg ccctccgaag
    11461 aggtcgtgaa gaagatgaag aactactggc ggcagctgct gaacgccaag ctgattaccc
    11521 agagaaagtt cgacaatctg accaaggccg agagaggcgg cctgagcgaa ctggataagg
    11581 ccggcttcat caagagacag ctggtggaaa cccggcagat cacaaagcac gtggcacaga
    11641 tcctggactc ccggatgaac actaagtacg acgagaatga caagctgatc cgggaagtga
    11701 aagtgatcac cctgaagtcc aagctggtgt ccgatttccg gaaggatttc cagttttaca
    11761 aagtgcgcga gatcaacaac taccaccacg cccacgacgc ctacctgaac gccgtcgtgg
    11821 gaaccgccct gatcaaaaag taccctaagc tggaaagcga gttcgtgtac ggcgactaca
    11881 aggtgtacga cgtgcggaag atgatcgcca agagcgagca ggaaatcggc aaggctaccg
    11941 ccaagtactt cttctacagc aacatcatga actttttcaa gaccgagatt accctggcca
    12001 acggcgagat ccggaagcgg cctctgatcg agacaaacgg cgaaaccggg gagatcgtgt
    12061 gggataaggg ccgggatttt gccaccgtgc ggaaagtgct gagcatgccc caagtgaata
    12121 tcgtgaaaaa gaccgaggtg cagacaggcg gcttcagcaa agagtctatc ctgcccaaga
    12181 ggaacagcga taagctgatc gccagaaaga aggactggga ccctaagaag tacggcggct
    12241 tcgacagccc caccgtggcc tattctgtgc tggtggtggc caaagtggaa aagggcaagt
    12301 ccaagaaact gaagagtgtg aaagagctgc tggggatcac catcatggaa agaagcagct
    12361 tcgagaagaa tcccatcgac tttctggaag ccaagggcta caaagaagtg aaaaaggacc
    12421 tgatcatcaa gctgcctaag tactccctgt tcgagctgga aaacggccgg aagagaatgc
    12481 tggcctctgc cggcgaactg cagaagggaa acgaactggc cctgccctcc aaatatgtga
    12541 acttcctgta cctggccagc cactatgaga agctgaaggg ctcccccgag gataatgagc
    12601 agaaacagct gtttgtggaa cagcacaagc actacctgga cgagatcatc gagcagatca
    12661 gcgagttctc caagagagtg atcctggccg acgctaatct ggacaaagtg ctgtccgcct
    12721 acaacaagca ccgggataag cccatcagag agcaggccga gaatatcatc cacctgttta
    12781 ccctgaccaa tctgggagcc cctgccgcct tcaagtactt tgacaccacc atcgaccgga
    12841 agaggtacac cagcaccaaa gaggtgctgg acgccaccct gatccaccag agcatcaccg
    12901 gcctgtacga gacacggatc gacctgtctc agctgggagg cgacaagcga cctgccgcca
    12961 caaagaaggc tggacaggct aagaagaaga aagattacaa agacgatgac gataagggat
    13021 ccggcgcaac aaacttctct ctgctgaaac aagccggaga tgtcgaagag aatcctggac
    13081 cgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg
    13141 gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag
    13201 cgcaggggcg cc
//
