LOCUS       sequence-413634-       13013 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  knock out BAX in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13013)
  AUTHORS   Liu X, Nie L, Zhang Y, Yan Y, Wang C, Colic M, Olszewski K, Horbath 
            A, Chen X, Lei G, Mao C, Wu S, Zhuang L, Poyurovsky MV, James You M,
            Hart T, Billadeau DD, Chen J, Gan B
  TITLE     Actin cytoskeleton vulnerability to disulfide stress mediates 
            disulfidptosis.
  JOURNAL   Nat Cell Biol. 2023 Mar;25(3):404-414. doi: 
            10.1038/s41556-023-01091-2. Epub 2023 Feb 6.
  PUBMED    36747082
REFERENCE   2  (bases 1 to 13013)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1038/s41556-023-01091-2"; journalName: "Nat Cell Biol"; date: 
            "2023-03"; volume: "25"; issue: "3"; pages: "404-414"
COMMENT     Sequence Label: lentiCRISPR v2-sgBAX-2
FEATURES             Location/Qualifiers
     source          1..13013
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        3..201
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     156..176
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             219..399
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    446..571
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1064..1297
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1482..1526
                     /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    1824..1941
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        1992..2240
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1992..2012
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2163..2182
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2262..2337
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        2399..2610
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      2629..2638
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2635..6738
                     /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             2635..6738
                     /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             6739..6786
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             6787..6810
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             6820..6876
                     /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             6877..7473
                     /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     7370..7390
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    7489..8077
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7542..7562)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(7960..7971)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             8149..8382
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(8408..8425)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    8414..8638
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      8684..9112
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8771..8790)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     8981..9002
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(9121..9141)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        9126..9455
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      9306..9441
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     9368..9387
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        9503..9550
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             9569..9943
                     /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    10073..10194
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(10110..10129)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     10164..10183
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(10243..10259)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(10243..10259)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(10256..10278)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    10267..10283
                     /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(10291..10297,10298..10315,10316..10321))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    10336..10357
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(10474..10491)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(10645..11233)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(10725..10744)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(11404..12195,12196..12264))
                     /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     12027..12046
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(12265..12369)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(12444..12463)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        join(12635..13013,1)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag cggtttgact cacggggatt
       61 tccaagtctc caccccattg acgtcaatgg gagtttgttt tggcaccaaa atcaacggga
      121 ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa atgggcggta ggcgtgtacg
      181 gtgggaggtc tatataagca gcgcgttttg cctgtactgg gtctctctgg ttagaccaga
      241 tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct
      301 tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat
      361 ccctcagacc cttttagtca gtgtggaaaa tctctagcag tggcgcccga acagggactt
      421 gaaagcgaaa gggaaaccag aggagctctc tcgacgcagg actcggcttg ctgaagcgcg
      481 cacggcaaga ggcgaggggc ggcgactggt gagtacgcca aaaattttga ctagcggagg
      541 ctagaaggag agagatgggt gcgagagcgt cagtattaag cgggggagaa ttagatcgcg
      601 atgggaaaaa attcggttaa ggccaggggg aaagaaaaaa tataaattaa aacatatagt
      661 atgggcaagc agggagctag aacgattcgc agttaatcct ggcctgttag aaacatcaga
      721 aggctgtaga caaatactgg gacagctaca accatccctt cagacaggat cagaagaact
      781 tagatcatta tataatacag tagcaaccct ctattgtgtg catcaaagga tagagataaa
      841 agacaccaag gaagctttag acaagataga ggaagagcaa aacaaaagta agaccaccgc
      901 acagcaagcg gccgctgatc ttcagacctg gaggaggaga tatgagggac aattggagaa
      961 gtgaattata taaatataaa gtagtaaaaa ttgaaccatt aggagtagca cccaccaagg
     1021 caaagagaag agtggtgcag agagaaaaaa gagcagtggg aataggagct ttgttccttg
     1081 ggttcttggg agcagcagga agcactatgg gcgcagcgtc aatgacgctg acggtacagg
     1141 ccagacaatt attgtctggt atagtgcagc agcagaacaa tttgctgagg gctattgagg
     1201 cgcaacagca tctgttgcaa ctcacagtct ggggcatcaa gcagctccag gcaagaatcc
     1261 tggctgtgga aagataccta aaggatcaac agctcctggg gatttggggt tgctctggaa
     1321 aactcatttg caccactgct gtgccttgga atgctagttg gagtaataaa tctctggaac
     1381 agatttggaa tcacacgacc tggatggagt gggacagaga aattaacaat tacacaagct
     1441 taatacactc cttaattgaa gaatcgcaaa accagcaaga aaagaatgaa caagaattat
     1501 tggaattaga taaatgggca agtttgtgga attggtttaa cataacaaat tggctgtggt
     1561 atataaaatt attcataatg atagtaggag gcttggtagg tttaagaata gtttttgctg
     1621 tactttctat agtgaataga gttaggcagg gatattcacc attatcgttt cagacccacc
     1681 tcccaacccc gaggggaccc gacaggcccg aaggaataga agaagaaggt ggagagagag
     1741 acagagacag atccattcga ttagtgaacg gatcggcact gcgtgcgcca attctgcaga
     1801 caaatggcag tattcatcca caattttaaa agaaaagggg ggattggggg gtacagtgca
     1861 ggggaaagaa tagtagacat aatagcaaca gacatacaaa ctaaagaatt acaaaaacaa
     1921 attacaaaaa ttcaaaattt tcgggtttat tacagggaca gcagagatcc agtttggtta
     1981 attaaggtac cgagggccta tttcccatga ttccttcata tttgcatata cgatacaagg
     2041 ctgttagaga gataattaga attaatttga ctgtaaacac aaagatatta gtacaaaata
     2101 cgtgacgtag aaagtaataa tttcttgggt agtttgcagt tttaaaatta tgttttaaaa
     2161 tggactatca tatgcttacc gtaacttgaa agtatttcga tttcttggct ttatatatct
     2221 tgtggaaagg acgaaacacc gcgagtgtct caagcgcatc ggttttagag ctagaaatag
     2281 caagttaaaa taaggctagt ccgttatcaa cttgaaaaag tggcaccgag tcggtgcttt
     2341 tttgaattcg ctagctaggt cttgaaagga gtgggaattg gctccggtgc ccgtcagtgg
     2401 gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg caattgatcc
     2461 ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta ctggctccgc
     2521 ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt gaacgttctt
     2581 tttcgcaacg ggtttgccgc cagaacacag gaccggttct agagcgctgc caccatggac
     2641 aagaagtaca gcatcggcct ggacatcggc accaactctg tgggctgggc cgtgatcacc
     2701 gacgagtaca aggtgcccag caagaaattc aaggtgctgg gcaacaccga ccggcacagc
     2761 atcaagaaga acctgatcgg agccctgctg ttcgacagcg gcgaaacagc cgaggccacc
     2821 cggctgaaga gaaccgccag aagaagatac accagacgga agaaccggat ctgctatctg
     2881 caagagatct tcagcaacga gatggccaag gtggacgaca gcttcttcca cagactggaa
     2941 gagtccttcc tggtggaaga ggataagaag cacgagcggc accccatctt cggcaacatc
     3001 gtggacgagg tggcctacca cgagaagtac cccaccatct accacctgag aaagaaactg
     3061 gtggacagca ccgacaaggc cgacctgcgg ctgatctatc tggccctggc ccacatgatc
     3121 aagttccggg gccacttcct gatcgagggc gacctgaacc ccgacaacag cgacgtggac
     3181 aagctgttca tccagctggt gcagacctac aaccagctgt tcgaggaaaa ccccatcaac
     3241 gccagcggcg tggacgccaa ggccatcctg tctgccagac tgagcaagag cagacggctg
     3301 gaaaatctga tcgcccagct gcccggcgag aagaagaatg gcctgttcgg aaacctgatt
     3361 gccctgagcc tgggcctgac ccccaacttc aagagcaact tcgacctggc cgaggatgcc
     3421 aaactgcagc tgagcaagga cacctacgac gacgacctgg acaacctgct ggcccagatc
     3481 ggcgaccagt acgccgacct gtttctggcc gccaagaacc tgtccgacgc catcctgctg
     3541 agcgacatcc tgagagtgaa caccgagatc accaaggccc ccctgagcgc ctctatgatc
     3601 aagagatacg acgagcacca ccaggacctg accctgctga aagctctcgt gcggcagcag
     3661 ctgcctgaga agtacaaaga gattttcttc gaccagagca agaacggcta cgccggctac
     3721 attgacggcg gagccagcca ggaagagttc tacaagttca tcaagcccat cctggaaaag
     3781 atggacggca ccgaggaact gctcgtgaag ctgaacagag aggacctgct gcggaagcag
     3841 cggaccttcg acaacggcag catcccccac cagatccacc tgggagagct gcacgccatt
     3901 ctgcggcggc aggaagattt ttacccattc ctgaaggaca accgggaaaa gatcgagaag
     3961 atcctgacct tccgcatccc ctactacgtg ggccctctgg ccaggggaaa cagcagattc
     4021 gcctggatga ccagaaagag cgaggaaacc atcaccccct ggaacttcga ggaagtggtg
     4081 gacaagggcg cttccgccca gagcttcatc gagcggatga ccaacttcga taagaacctg
     4141 cccaacgaga aggtgctgcc caagcacagc ctgctgtacg agtacttcac cgtgtataac
     4201 gagctgacca aagtgaaata cgtgaccgag ggaatgagaa agcccgcctt cctgagcggc
     4261 gagcagaaaa aggccatcgt ggacctgctg ttcaagacca accggaaagt gaccgtgaag
     4321 cagctgaaag aggactactt caagaaaatc gagtgcttcg actccgtgga aatctccggc
     4381 gtggaagatc ggttcaacgc ctccctgggc acataccacg atctgctgaa aattatcaag
     4441 gacaaggact tcctggacaa tgaggaaaac gaggacattc tggaagatat cgtgctgacc
     4501 ctgacactgt ttgaggacag agagatgatc gaggaacggc tgaaaaccta tgcccacctg
     4561 ttcgacgaca aagtgatgaa gcagctgaag cggcggagat acaccggctg gggcaggctg
     4621 agccggaagc tgatcaacgg catccgggac aagcagtccg gcaagacaat cctggatttc
     4681 ctgaagtccg acggcttcgc caacagaaac ttcatgcagc tgatccacga cgacagcctg
     4741 acctttaaag aggacatcca gaaagcccag gtgtccggcc agggcgatag cctgcacgag
     4801 cacattgcca atctggccgg cagccccgcc attaagaagg gcatcctgca gacagtgaag
     4861 gtggtggacg agctcgtgaa agtgatgggc cggcacaagc ccgagaacat cgtgatcgaa
     4921 atggccagag agaaccagac cacccagaag ggacagaaga acagccgcga gagaatgaag
     4981 cggatcgaag agggcatcaa agagctgggc agccagatcc tgaaagaaca ccccgtggaa
     5041 aacacccagc tgcagaacga gaagctgtac ctgtactacc tgcagaatgg gcgggatatg
     5101 tacgtggacc aggaactgga catcaaccgg ctgtccgact acgatgtgga ccatatcgtg
     5161 cctcagagct ttctgaagga cgactccatc gacaacaagg tgctgaccag aagcgacaag
     5221 aaccggggca agagcgacaa cgtgccctcc gaagaggtcg tgaagaagat gaagaactac
     5281 tggcggcagc tgctgaacgc caagctgatt acccagagaa agttcgacaa tctgaccaag
     5341 gccgagagag gcggcctgag cgaactggat aaggccggct tcatcaagag acagctggtg
     5401 gaaacccggc agatcacaaa gcacgtggca cagatcctgg actcccggat gaacactaag
     5461 tacgacgaga atgacaagct gatccgggaa gtgaaagtga tcaccctgaa gtccaagctg
     5521 gtgtccgatt tccggaagga tttccagttt tacaaagtgc gcgagatcaa caactaccac
     5581 cacgcccacg acgcctacct gaacgccgtc gtgggaaccg ccctgatcaa aaagtaccct
     5641 aagctggaaa gcgagttcgt gtacggcgac tacaaggtgt acgacgtgcg gaagatgatc
     5701 gccaagagcg agcaggaaat cggcaaggct accgccaagt acttcttcta cagcaacatc
     5761 atgaactttt tcaagaccga gattaccctg gccaacggcg agatccggaa gcggcctctg
     5821 atcgagacaa acggcgaaac cggggagatc gtgtgggata agggccggga ttttgccacc
     5881 gtgcggaaag tgctgagcat gccccaagtg aatatcgtga aaaagaccga ggtgcagaca
     5941 ggcggcttca gcaaagagtc tatcctgccc aagaggaaca gcgataagct gatcgccaga
     6001 aagaaggact gggaccctaa gaagtacggc ggcttcgaca gccccaccgt ggcctattct
     6061 gtgctggtgg tggccaaagt ggaaaagggc aagtccaaga aactgaagag tgtgaaagag
     6121 ctgctgggga tcaccatcat ggaaagaagc agcttcgaga agaatcccat cgactttctg
     6181 gaagccaagg gctacaaaga agtgaaaaag gacctgatca tcaagctgcc taagtactcc
     6241 ctgttcgagc tggaaaacgg ccggaagaga atgctggcct ctgccggcga actgcagaag
     6301 ggaaacgaac tggccctgcc ctccaaatat gtgaacttcc tgtacctggc cagccactat
     6361 gagaagctga agggctcccc cgaggataat gagcagaaac agctgtttgt ggaacagcac
     6421 aagcactacc tggacgagat catcgagcag atcagcgagt tctccaagag agtgatcctg
     6481 gccgacgcta atctggacaa agtgctgtcc gcctacaaca agcaccggga taagcccatc
     6541 agagagcagg ccgagaatat catccacctg tttaccctga ccaatctggg agcccctgcc
     6601 gccttcaagt actttgacac caccatcgac cggaagaggt acaccagcac caaagaggtg
     6661 ctggacgcca ccctgatcca ccagagcatc accggcctgt acgagacacg gatcgacctg
     6721 tctcagctgg gaggcgacaa gcgacctgcc gccacaaaga aggctggaca ggctaagaag
     6781 aagaaagatt acaaagacga tgacgataag ggatccggcg caacaaactt ctctctgctg
     6841 aaacaagccg gagatgtcga agagaatcct ggaccgaccg agtacaagcc cacggtgcgc
     6901 ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac
     6961 taccccgcca cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg
     7021 caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac
     7081 ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc
     7141 gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg
     7201 gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcgga
     7261 gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag
     7321 gcggccgagc gcgccggggt gcccgccttc ctggagacct ccgcgccccg caacctcccc
     7381 ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc
     7441 acctggtgca tgacccgcaa gcccggtgcc tgaacgcgtt aagtcgacaa tcaacctctg
     7501 gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta
     7561 tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt
     7621 ttctcctcct tgtataaatc ctggttgctg tctctttatg aggagttgtg gcccgttgtc
     7681 aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg ttggggcatt
     7741 gccaccacct gtcagctcct ttccgggact ttcgctttcc ccctccctat tgccacggcg
     7801 gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac
     7861 aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc ctgtgttgcc
     7921 acctggattc tgcgcgggac gtccttctgc tacgtccctt cggccctcaa tccagcggac
     7981 cttccttccc gcggcctgct gccggctctg cggcctcttc cgcgtcttcg ccttcgccct
     8041 cagacgagtc ggatctccct ttgggccgcc tccccgcgtc gactttaaga ccaatgactt
     8101 acaaggcagc tgtagatctt agccactttt taaaagaaaa ggggggactg gaagggctaa
     8161 ttcactccca acgaagacaa gatctgcttt ttgcttgtac tgggtctctc tggttagacc
     8221 agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa
     8281 gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga
     8341 gatccctcag acccttttag tcagtgtgga aaatctctag cagggcccgt ttaaacccgc
     8401 tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg
     8461 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt
     8521 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc
     8581 aagggggagg attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct
     8641 tctgaggcgg aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc
     8701 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     8761 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     8821 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     8881 gaccccaaaa aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     8941 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     9001 ggaacaacac tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt
     9061 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt
     9121 ggaatgtgtg tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     9181 aaagcatgca tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag
     9241 gcagaagtat gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc
     9301 cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa
     9361 ttttttttat ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt
     9421 gaggaggctt ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca
     9481 ttttcggatc tgatcagcac gtgttgacaa ttaatcatcg gcatagtata tcggcatagt
     9541 ataatacgac aaggtgagga actaaaccat ggccaagttg accagtgccg ttccggtgct
     9601 caccgcgcgc gacgtcgccg gagcggtcga gttctggacc gaccggctcg ggttctcccg
     9661 ggacttcgtg gaggacgact tcgccggtgt ggtccgggac gacgtgaccc tgttcatcag
     9721 cgcggtccag gaccaggtgg tgccggacaa caccctggcc tgggtgtggg tgcgcggcct
     9781 ggacgagctg tacgccgagt ggtcggaggt cgtgtccacg aacttccggg acgcctccgg
     9841 gccggccatg accgagatcg gcgagcagcc gtgggggcgg gagttcgccc tgcgcgaccc
     9901 ggccggcaac tgcgtgcact tcgtggccga ggagcaggac tgacacgtgc tacgagattt
     9961 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
    10021 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt
    10081 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
    10141 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
    10201 ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt
    10261 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
    10321 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
    10381 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
    10441 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
    10501 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
    10561 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
    10621 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
    10681 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
    10741 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
    10801 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
    10861 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
    10921 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
    10981 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
    11041 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
    11101 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
    11161 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
    11221 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
    11281 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
    11341 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
    11401 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
    11461 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
    11521 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
    11581 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
    11641 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
    11701 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
    11761 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
    11821 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
    11881 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
    11941 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
    12001 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
    12061 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
    12121 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
    12181 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
    12241 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
    12301 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
    12361 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatctccc
    12421 gatcccctat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat
    12481 ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg cgagcaaaat ttaagctaca
    12541 acaaggcaag gcttgaccga caattgcatg aagaatctgc ttagggttag gcgttttgcg
    12601 ctgcttcgcg atgtacgggc cagatatacg cgttgacatt gattattgac tagttattaa
    12661 tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg cgttacataa
    12721 cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt gacgtcaata
    12781 atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag
    12841 tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc
    12901 cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta
    12961 tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac cat
//
