LOCUS       sequence-340297-       13346 bp    DNA     circular SYN 15-APR-2025
DEFINITION  A knock-out vector for the dog Egf.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13346)
  AUTHORS   Lin S, Hirayama D, Maryu G, Matsuda K, Hino N, Deguchi E, Aoki K, 
            Iwamoto R, Terai K, Matsuda M
  TITLE     Redundant roles of EGFR ligands in the ERK activation waves during 
            collective cell migration.
  JOURNAL   Life Sci Alliance. 2021 Oct 19;5(1). pii: 5/1/e202101206. doi: 
            10.26508/lsa.202101206. Print 2022 Jan.
  PUBMED    34667080
REFERENCE   2  (bases 1 to 13346)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Life Sci 
            Alliance."; date: "2021-10-19"; volume: "5(1). pii: 5/1/e202101206. 
            doi"; pages: " 10.26508/lsa.202101206. Print 2022 Jan"
COMMENT     Sequence Label: lentiCRISPRv2-bleo-dEgf
FEATURES             Location/Qualifiers
     source          1..13346
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(150..169)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        341..720
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        722..920
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     875..895
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             938..1118
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1165..1290
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1783..2016
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2201..2245
                     /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    2543..2660
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2711..2959
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2711..2731
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2882..2901
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2981..3056
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        3118..3329
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      3348..3357
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3354..7457
                     /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             3354..7457
                     /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             7458..7505
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7506..7529
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     misc_feature    7599..8148
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(7743..7760)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     7970..7989
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             8151..8525
                     /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"
     misc_feature    8541..9129
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8594..8614)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(9012..9023)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             9201..9434
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(9460..9477)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9466..9690
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      9736..10164
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9823..9842)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10033..10054
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10173..10193)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10178..10507
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      10358..10493
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10420..10439
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10555..10602
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10621..10995
                     /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    11125..11246
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11162..11181)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11216..11235
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11295..11311)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11295..11311)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11308..11330)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11319..11335
                     /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(11343..11349,11350..11367,11368..11373))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11388..11409
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11526..11543)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11697..12285)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11777..11796)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12456..13247,13248..13316))
                     /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     13079..13098
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(13317..13346,1..75))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
       61 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gacgtcgacg gatcgggaga
      121 tctcccgatc ccctatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc
      181 cagtatctgc tccctgcttg tgtgttggag gtcgctgagt agtgcgcgag caaaatttaa
      241 gctacaacaa ggcaaggctt gaccgacaat tgcatgaaga atctgcttag ggttaggcgt
      301 tttgcgctgc ttcgcgatgt acgggccaga tatacgcgtt gacattgatt attgactagt
      361 tattaatagt aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt
      421 acataactta cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg
      481 tcaataatga cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg
      541 gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt
      601 acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg
      661 accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg
      721 gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc acggggattt
      781 ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac
      841 tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag gcgtgtacgg
      901 tgggaggtct atataagcag cgcgttttgc ctgtactggg tctctctggt tagaccagat
      961 ctgagcctgg gagctctctg gctaactagg gaacccactg cttaagcctc aataaagctt
     1021 gccttgagtg cttcaagtag tgtgtgcccg tctgttgtgt gactctggta actagagatc
     1081 cctcagaccc ttttagtcag tgtggaaaat ctctagcagt ggcgcccgaa cagggacttg
     1141 aaagcgaaag ggaaaccaga ggagctctct cgacgcagga ctcggcttgc tgaagcgcgc
     1201 acggcaagag gcgaggggcg gcgactggtg agtacgccaa aaattttgac tagcggaggc
     1261 tagaaggaga gagatgggtg cgagagcgtc agtattaagc gggggagaat tagatcgcga
     1321 tgggaaaaaa ttcggttaag gccaggggga aagaaaaaat ataaattaaa acatatagta
     1381 tgggcaagca gggagctaga acgattcgca gttaatcctg gcctgttaga aacatcagaa
     1441 ggctgtagac aaatactggg acagctacaa ccatcccttc agacaggatc agaagaactt
     1501 agatcattat ataatacagt agcaaccctc tattgtgtgc atcaaaggat agagataaaa
     1561 gacaccaagg aagctttaga caagatagag gaagagcaaa acaaaagtaa gaccaccgca
     1621 cagcaagcgg ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag
     1681 tgaattatat aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc
     1741 aaagagaaga gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg
     1801 gttcttggga gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc
     1861 cagacaatta ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc
     1921 gcaacagcat ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct
     1981 ggctgtggaa agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa
     2041 actcatttgc accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca
     2101 gatttggaat cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt
     2161 aatacactcc ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt
     2221 ggaattagat aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta
     2281 tataaaatta ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt
     2341 actttctata gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct
     2401 cccaaccccg aggggacccg acaggcccga aggaatagaa gaagaaggtg gagagagaga
     2461 cagagacaga tccattcgat tagtgaacgg atcggcactg cgtgcgccaa ttctgcagac
     2521 aaatggcagt attcatccac aattttaaaa gaaaaggggg gattgggggg tacagtgcag
     2581 gggaaagaat agtagacata atagcaacag acatacaaac taaagaatta caaaaacaaa
     2641 ttacaaaaat tcaaaatttt cgggtttatt acagggacag cagagatcca gtttggttaa
     2701 ttaaggtacc gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc
     2761 tgttagagag ataattagaa ttaatttgac tgtaaacaca aagatattag tacaaaatac
     2821 gtgacgtaga aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat
     2881 ggactatcat atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt
     2941 gtggaaagga cgaaacaccg accggccaca gaatgataag gttttagagc tagaaatagc
     3001 aagttaaaat aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt
     3061 ttgaattcgc tagctaggtc ttgaaaggag tgggaattgg ctccggtgcc cgtcagtggg
     3121 cagagcgcac atcgcccaca gtccccgaga agttgggggg aggggtcggc aattgatccg
     3181 gtgcctagag aaggtggcgc ggggtaaact gggaaagtga tgtcgtgtac tggctccgcc
     3241 tttttcccga gggtggggga gaaccgtata taagtgcagt agtcgccgtg aacgttcttt
     3301 ttcgcaacgg gtttgccgcc agaacacagg accggttcta gagcgctgcc accatggaca
     3361 agaagtacag catcggcctg gacatcggca ccaactctgt gggctgggcc gtgatcaccg
     3421 acgagtacaa ggtgcccagc aagaaattca aggtgctggg caacaccgac cggcacagca
     3481 tcaagaagaa cctgatcgga gccctgctgt tcgacagcgg cgaaacagcc gaggccaccc
     3541 ggctgaagag aaccgccaga agaagataca ccagacggaa gaaccggatc tgctatctgc
     3601 aagagatctt cagcaacgag atggccaagg tggacgacag cttcttccac agactggaag
     3661 agtccttcct ggtggaagag gataagaagc acgagcggca ccccatcttc ggcaacatcg
     3721 tggacgaggt ggcctaccac gagaagtacc ccaccatcta ccacctgaga aagaaactgg
     3781 tggacagcac cgacaaggcc gacctgcggc tgatctatct ggccctggcc cacatgatca
     3841 agttccgggg ccacttcctg atcgagggcg acctgaaccc cgacaacagc gacgtggaca
     3901 agctgttcat ccagctggtg cagacctaca accagctgtt cgaggaaaac cccatcaacg
     3961 ccagcggcgt ggacgccaag gccatcctgt ctgccagact gagcaagagc agacggctgg
     4021 aaaatctgat cgcccagctg cccggcgaga agaagaatgg cctgttcgga aacctgattg
     4081 ccctgagcct gggcctgacc cccaacttca agagcaactt cgacctggcc gaggatgcca
     4141 aactgcagct gagcaaggac acctacgacg acgacctgga caacctgctg gcccagatcg
     4201 gcgaccagta cgccgacctg tttctggccg ccaagaacct gtccgacgcc atcctgctga
     4261 gcgacatcct gagagtgaac accgagatca ccaaggcccc cctgagcgcc tctatgatca
     4321 agagatacga cgagcaccac caggacctga ccctgctgaa agctctcgtg cggcagcagc
     4381 tgcctgagaa gtacaaagag attttcttcg accagagcaa gaacggctac gccggctaca
     4441 ttgacggcgg agccagccag gaagagttct acaagttcat caagcccatc ctggaaaaga
     4501 tggacggcac cgaggaactg ctcgtgaagc tgaacagaga ggacctgctg cggaagcagc
     4561 ggaccttcga caacggcagc atcccccacc agatccacct gggagagctg cacgccattc
     4621 tgcggcggca ggaagatttt tacccattcc tgaaggacaa ccgggaaaag atcgagaaga
     4681 tcctgacctt ccgcatcccc tactacgtgg gccctctggc caggggaaac agcagattcg
     4741 cctggatgac cagaaagagc gaggaaacca tcaccccctg gaacttcgag gaagtggtgg
     4801 acaagggcgc ttccgcccag agcttcatcg agcggatgac caacttcgat aagaacctgc
     4861 ccaacgagaa ggtgctgccc aagcacagcc tgctgtacga gtacttcacc gtgtataacg
     4921 agctgaccaa agtgaaatac gtgaccgagg gaatgagaaa gcccgccttc ctgagcggcg
     4981 agcagaaaaa ggccatcgtg gacctgctgt tcaagaccaa ccggaaagtg accgtgaagc
     5041 agctgaaaga ggactacttc aagaaaatcg agtgcttcga ctccgtggaa atctccggcg
     5101 tggaagatcg gttcaacgcc tccctgggca cataccacga tctgctgaaa attatcaagg
     5161 acaaggactt cctggacaat gaggaaaacg aggacattct ggaagatatc gtgctgaccc
     5221 tgacactgtt tgaggacaga gagatgatcg aggaacggct gaaaacctat gcccacctgt
     5281 tcgacgacaa agtgatgaag cagctgaagc ggcggagata caccggctgg ggcaggctga
     5341 gccggaagct gatcaacggc atccgggaca agcagtccgg caagacaatc ctggatttcc
     5401 tgaagtccga cggcttcgcc aacagaaact tcatgcagct gatccacgac gacagcctga
     5461 cctttaaaga ggacatccag aaagcccagg tgtccggcca gggcgatagc ctgcacgagc
     5521 acattgccaa tctggccggc agccccgcca ttaagaaggg catcctgcag acagtgaagg
     5581 tggtggacga gctcgtgaaa gtgatgggcc ggcacaagcc cgagaacatc gtgatcgaaa
     5641 tggccagaga gaaccagacc acccagaagg gacagaagaa cagccgcgag agaatgaagc
     5701 ggatcgaaga gggcatcaaa gagctgggca gccagatcct gaaagaacac cccgtggaaa
     5761 acacccagct gcagaacgag aagctgtacc tgtactacct gcagaatggg cgggatatgt
     5821 acgtggacca ggaactggac atcaaccggc tgtccgacta cgatgtggac catatcgtgc
     5881 ctcagagctt tctgaaggac gactccatcg acaacaaggt gctgaccaga agcgacaaga
     5941 accggggcaa gagcgacaac gtgccctccg aagaggtcgt gaagaagatg aagaactact
     6001 ggcggcagct gctgaacgcc aagctgatta cccagagaaa gttcgacaat ctgaccaagg
     6061 ccgagagagg cggcctgagc gaactggata aggccggctt catcaagaga cagctggtgg
     6121 aaacccggca gatcacaaag cacgtggcac agatcctgga ctcccggatg aacactaagt
     6181 acgacgagaa tgacaagctg atccgggaag tgaaagtgat caccctgaag tccaagctgg
     6241 tgtccgattt ccggaaggat ttccagtttt acaaagtgcg cgagatcaac aactaccacc
     6301 acgcccacga cgcctacctg aacgccgtcg tgggaaccgc cctgatcaaa aagtacccta
     6361 agctggaaag cgagttcgtg tacggcgact acaaggtgta cgacgtgcgg aagatgatcg
     6421 ccaagagcga gcaggaaatc ggcaaggcta ccgccaagta cttcttctac agcaacatca
     6481 tgaacttttt caagaccgag attaccctgg ccaacggcga gatccggaag cggcctctga
     6541 tcgagacaaa cggcgaaacc ggggagatcg tgtgggataa gggccgggat tttgccaccg
     6601 tgcggaaagt gctgagcatg ccccaagtga atatcgtgaa aaagaccgag gtgcagacag
     6661 gcggcttcag caaagagtct atcctgccca agaggaacag cgataagctg atcgccagaa
     6721 agaaggactg ggaccctaag aagtacggcg gcttcgacag ccccaccgtg gcctattctg
     6781 tgctggtggt ggccaaagtg gaaaagggca agtccaagaa actgaagagt gtgaaagagc
     6841 tgctggggat caccatcatg gaaagaagca gcttcgagaa gaatcccatc gactttctgg
     6901 aagccaaggg ctacaaagaa gtgaaaaagg acctgatcat caagctgcct aagtactccc
     6961 tgttcgagct ggaaaacggc cggaagagaa tgctggcctc tgccggcgaa ctgcagaagg
     7021 gaaacgaact ggccctgccc tccaaatatg tgaacttcct gtacctggcc agccactatg
     7081 agaagctgaa gggctccccc gaggataatg agcagaaaca gctgtttgtg gaacagcaca
     7141 agcactacct ggacgagatc atcgagcaga tcagcgagtt ctccaagaga gtgatcctgg
     7201 ccgacgctaa tctggacaaa gtgctgtccg cctacaacaa gcaccgggat aagcccatca
     7261 gagagcaggc cgagaatatc atccacctgt ttaccctgac caatctggga gcccctgccg
     7321 ccttcaagta ctttgacacc accatcgacc ggaagaggta caccagcacc aaagaggtgc
     7381 tggacgccac cctgatccac cagagcatca ccggcctgta cgagacacgg atcgacctgt
     7441 ctcagctggg aggcgacaag cgacctgccg ccacaaagaa ggctggacag gctaagaaga
     7501 agaaagatta caaagacgat gacgataagg gatccgttaa ctaacttaag ctagcaacgg
     7561 tttccctcta gcgggatcaa ttccgccccc cccccctaac gttactggcc gaagccgctt
     7621 ggaataaggc cggtgtgcgt ttgtctatat gttattttcc accatattgc cgtcttttgg
     7681 caatgtgagg gcccggaaac ctggccctgt cttcttgacg agcattccta ggggtctttc
     7741 ccctctcgcc aaaggaatgc aaggtctgtt gaatgtcgtg aaggaagcag ttcctctgga
     7801 agcttcttga agacaaacaa cgtctgtagc gaccctttgc aggcagcgga accccccacc
     7861 tggcgacagg tgcctctgcg gccaaaagcc acgtgtataa gatacacctg caaaggcggc
     7921 acaaccccag tgccacgttg tgagttggat agttgtggaa agagtcaaat ggctctcctc
     7981 aagcgtattc aacaaggggc tgaaggatgc ccagaaggta ccccattgta tgggatctga
     8041 tctggggcct cggtgcacat gctttacatg tgtttagtcg aggttaaaaa aacgtctagg
     8101 ccccccgaac cacggggacg tggttttcct ttgaaaaaca cgataatacc atggccaagt
     8161 tgaccagtgc cgttccggtg ctcaccgcgc gcgacgtcgc cggagcggtc gagttctgga
     8221 ccgaccggct cgggttctcc cgggacttcg tggaggacga cttcgccggt gtggtccggg
     8281 acgacgtgac cctgttcatc agcgcggtcc aggaccaggt ggtgccggac aacaccctgg
     8341 cctgggtgtg ggtgcgcggc ctggacgagc tgtacgccga gtggtcggag gtcgtgtcca
     8401 cgaacttccg ggacgcctcc gggccggcca tgaccgagat cggcgagcag ccgtgggggc
     8461 gggagttcgc cctgcgcgac ccggccggca actgcgtgca cttcgtggcc gaggagcagg
     8521 actgaacgcg ttaagtcgac aatcaacctc tggattacaa aatttgtgaa agattgactg
     8581 gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt
     8641 atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc
     8701 tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt
     8761 ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga
     8821 ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct
     8881 gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat
     8941 cgtcctttcc ttggctgctc gcctgtgttg ccacctggat tctgcgcggg acgtccttct
     9001 gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc
     9061 tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg
     9121 cctccccgcg tcgactttaa gaccaatgac ttacaaggca gctgtagatc ttagccactt
     9181 tttaaaagaa aaggggggac tggaagggct aattcactcc caacgaagac aagatctgct
     9241 ttttgcttgt actgggtctc tctggttaga ccagatctga gcctgggagc tctctggcta
     9301 actagggaac ccactgctta agcctcaata aagcttgcct tgagtgcttc aagtagtgtg
     9361 tgcccgtctg ttgtgtgact ctggtaacta gagatccctc agaccctttt agtcagtgtg
     9421 gaaaatctct agcagggccc gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt
     9481 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     9541 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
     9601 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca
     9661 ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct
     9721 ctagggggta tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     9781 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     9841 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     9901 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     9961 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
    10021 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
    10081 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
    10141 tttaacaaaa atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa
    10201 gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac
    10261 caggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
    10321 ttagtcagca accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
    10381 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
    10441 cgcctctgcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt
    10501 ttgcaaaaag ctcccgggag cttgtatatc cattttcgga tctgatcagc acgtgttgac
    10561 aattaatcat cggcatagta tatcggcata gtataatacg acaaggtgag gaactaaacc
    10621 atggccaagt tgaccagtgc cgttccggtg ctcaccgcgc gcgacgtcgc cggagcggtc
    10681 gagttctgga ccgaccggct cgggttctcc cgggacttcg tggaggacga cttcgccggt
    10741 gtggtccggg acgacgtgac cctgttcatc agcgcggtcc aggaccaggt ggtgccggac
    10801 aacaccctgg cctgggtgtg ggtgcgcggc ctggacgagc tgtacgccga gtggtcggag
    10861 gtcgtgtcca cgaacttccg ggacgcctcc gggccggcca tgaccgagat cggcgagcag
    10921 ccgtgggggc gggagttcgc cctgcgcgac ccggccggca actgcgtgca cttcgtggcc
    10981 gaggagcagg actgacacgt gctacgagat ttcgattcca ccgccgcctt ctatgaaagg
    11041 ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg cggggatctc
    11101 atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg ttacaaataa
    11161 agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt
    11221 ttgtccaaac tcatcaatgt atcttatcat gtctgtatac cgtcgacctc tagctagagc
    11281 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
    11341 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
    11401 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
    11461 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
    11521 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct
    11581 cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg aaagaacatg
    11641 tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc
    11701 cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca gaggtggcga
    11761 aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct cgtgcgctct
    11821 cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc gggaagcgtg
    11881 gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag
    11941 ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc cggtaactat
    12001 cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc cactggtaac
    12061 aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg gtggcctaac
    12121 tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc agttaccttc
    12181 ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag cggtggtttt
    12241 tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc
    12301 ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat tttggtcatg
    12361 agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca
    12421 atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca
    12481 cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag
    12541 ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgagac
    12601 ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc
    12661 agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct
    12721 agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc
    12781 gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg
    12841 cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc
    12901 gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat
    12961 tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag
    13021 tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat
    13081 aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg
    13141 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
    13201 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
    13261 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
    13321 ttcctttttc aatattattg aagcat
//
