LOCUS       sequence-484728-       13112 bp    DNA     circular SYN 27-AUG-2025
DEFINITION  Sleeping Beauty XPRESSO vector for Cas9-EGFP expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13112)
  AUTHORS   Wexler Y, Grinstein H, Huber I, Glatstein S, Ghiringhelli M, Edri O,
            Landesberg M, Shiff D, Arbel G, Rosh I, Choudhary A, Stern S, 
            Gepstein L
  TITLE     XPRESSO: Rapid genetic engineering of human pluripotent stem cells 
            for durable overexpression using a modular anti-silencing vector.
  JOURNAL   Stem Cell Reports. 2025 Aug 19:102603. doi: 
            10.1016/j.stemcr.2025.102603.
  PUBMED    40845851
REFERENCE   2  (bases 1 to 13112)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 27, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Stem Cell 
            Reports. 2025 Aug 19:102603. doi: 10.1016/j.stemcr.2025.102603."
COMMENT     Sequence Label: pSB-XPRESSO-hSpCas9-EGFP
FEATURES             Location/Qualifiers
     source          1..13112
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    324..548
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     537..554
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     misc_feature    587..1175
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             693..704
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1102..1122
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(join(1203..1916,1917..1919))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             complement(join(1206..1916,1917..1919))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1248..1269)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1597..1616
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     CDS             complement(1920..1973)
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             complement(1989..2036)
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             complement(2037..6137)
                     /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="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             complement(2037..6137)
                     /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="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             complement(6162..6182)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     regulatory      6185..6194
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(6259..6278)
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     intron          6286..7302
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     complement(6340..6362)
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     enhancer        7580..7959
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     CDS             9130..9831
                     /codon_start=1
                     /product="dimeric variant of DsRed fluorescent protein 
                     (Shaner et al., 2004)"
                     /label=dTomato
                     /note="mammalian codon-optimized"
                     /translation="MLSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLYGMDELYK"
     primer_bind     complement(9263..9281)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     9531..9550
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     9725..9748
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     CDS             9841..9897
                     /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             9898..10497
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(9898..9917)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     10394..10414
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(10597..10616)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    10662..10717
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(10716..10735)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(11109..11126)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11280..11868)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11360..11379)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12039..12830,12831..12899))
                     /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     12662..12681
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(12900..13004)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggatccctat acagttgaag tcggaagttt acatacactt aagttggagt cattaaaact
       61 cgtttttcaa ctactccaca aatttcttgt taacaaacaa tagttttggc aagtcagtta
      121 ggacatctac tttgtgcatg acacaagtca tttttccaac aattgtttac agacagatta
      181 tttcacttat aattcactgt atcacaattc cagtgggtca gaagtttaca tacactaagt
      241 tcgactcctc tgcagaatgc ggcgatgttt cggtaagggg tccgctatct agtaggccca
      301 gctggttctt tccgcctcag aagccataga gcccaccgca tccccagcat gcctgctatt
      361 gtcttcccaa tcctccccct tgctgtcctg ccccacccca ccccccagaa tagaatgaca
      421 cctactcaga caatgcgatg caatttcctc attttattag gaaaggacag tgggagtggc
      481 accttccagg gtcaaggaag gcacggggga ggggcaaaca acagatggct ggcaactaga
      541 aggcacagtc gaggctgatc agcgagctca ggtcgacggt atcgatgcgg ggaggcggcc
      601 caaagggaga tccgactcgt ctgagggcga aggcgaagac gcggaagagg ccgcagagcc
      661 ggcagcaggc cgcgggaagg aaggtccgct ggattgaggg ccgaagggac gtagcagaag
      721 gacgtcccgc gcagaatcca ggtggcaaca caggcgagca gccaaggaaa ggacgatgat
      781 ttccccgaca acaccacgga attgtcagtg cccaacagcc gagcccctgt ccagcagcgg
      841 gcaaggcagg cggcgatgag ttccgccgtg gcaataggga gggggaaagc gaaagtcccg
      901 gaaaggagct gacaggtggt ggcaatgccc caaccagtgg gggttgcgtc agcaaacaca
      961 gtgcacacca cgccacgttg cctgacaacg ggccacaact cctcataaag agacagcaac
     1021 caggatttat acaaggagga gaaaatgaaa gccatacggg aagcaatagc atgatacaaa
     1081 ggcattaaag cagcgtatcc acatagcgta aaaggagcaa catagttaag aataccagtc
     1141 aatctttcac aaattttgta atccagaggt tgattatcga aagcttggcc tgacaggccg
     1201 tctcacttgt acagctcgtc catgccgaga gtgatcccgg cggcggtcac gaactccagc
     1261 aggaccatgt gatcgcgctt ctcgttgggg tctttgctca gggcggactg ggtgctcagg
     1321 tagtggttgt cgggcagcag cacggggccg tcgccgatgg gggtgttctg ctggtagtgg
     1381 tcggcgagct gcacgctgcc gtcctcgatg ttgtggcgga tcttgaagtt caccttgatg
     1441 ccgttcttct gcttgtcggc catgatatag acgttgtggc tgttgtagtt gtactccagc
     1501 ttgtgcccca ggatgttgcc gtcctccttg aagtcgatgc ccttcagctc gatgcggttc
     1561 accagggtgt cgccctcgaa cttcacctcg gcgcgggtct tgtagttgcc gtcgtccttg
     1621 aagaagatgg tgcgctcctg gacgtagcct tcgggcatgg cggacttgaa gaagtcgtgc
     1681 tgcttcatgt ggtcggggta gcggctgaag cactgcacgc cgtaggtcag ggtggtcacg
     1741 agggtgggcc agggcacggg cagcttgccg gtggtgcaga tgaacttcag ggtcagcttg
     1801 ccgtaggtgg catcgccctc gccctcgccg gacacgctga acttgtggcc gtttacgtcg
     1861 ccgtccagct cgaccaggat gggcaccacc ccggtgaaca gctcctcgcc cttgctcact
     1921 gggccaggat tctcctcgac gtcaccgcat gttagcagac ttcctctgcc ctctccactg
     1981 ccgaattcct ttttcttttt tgcctggccg gcctttttcg tggccgccgg ccttttgtcg
     2041 cctcccagct gagacaggtc gatccgtgtc tcgtacaggc cggtgatgct ctggtggatc
     2101 agggtggcgt ccagcacctc tttggtgctg gtgtacctct tccggtcgat ggtggtgtca
     2161 aagtacttga aggcggcagg ggctcccaga ttggtcaggg taaacaggtg gatgatattc
     2221 tcggcctgct ctctgatggg cttatcccgg tgcttgttgt aggcggacag cactttgtcc
     2281 agattagcgt cggccaggat cactctcttg gagaactcgc tgatctgctc gatgatctcg
     2341 tccaggtagt gcttgtgctg ttccacaaac agctgtttct gctcattatc ctcgggggag
     2401 cccttcagct tctcatagtg gctggccagg tacaggaagt tcacatattt ggagggcagg
     2461 gccagttcgt ttcccttctg cagttcgccg gcagaggcca gcattctctt ccggccgttt
     2521 tccagctcga acagggagta cttaggcagc ttgatgatca ggtccttttt cacttctttg
     2581 tagcccttgg cttccagaaa gtcgatggga ttcttctcga agctgcttct ttccatgatg
     2641 gtgatcccca gcagctcttt cacactcttc agtttcttgg acttgccctt ttccactttg
     2701 gccaccacca gcacagaata ggccacggtg gggctgtcga agccgccgta cttcttaggg
     2761 tcccagtcct tctttctggc gatcagctta tcgctgttcc tcttgggcag gatagactct
     2821 ttgctgaagc cgcctgtctg cacctcggtc tttttcacga tattcacttg gggcatgctc
     2881 agcactttcc gcacggtggc aaaatcccgg cccttatccc acacgatctc cccggtttcg
     2941 ccgtttgtct cgatcagagg ccgcttccgg atctcgccgt tggccagggt aatctcggtc
     3001 ttgaaaaagt tcatgatgtt gctgtagaag aagtacttgg cggtagcctt gccgatttcc
     3061 tgctcgctct tggcgatcat cttccgcacg tcgtacacct tgtagtcgcc gtacacgaac
     3121 tcgctttcca gcttagggta ctttttgatc agggcggttc ccacgacggc gttcaggtag
     3181 gcgtcgtggg cgtggtggta gttgttgatc tcgcgcactt tgtaaaactg gaaatccttc
     3241 cggaaatcgg acaccagctt ggacttcagg gtgatcactt tcacttcccg gatcagcttg
     3301 tcattctcgt cgtacttagt gttcatccgg gagtccagga tctgtgccac gtgctttgtg
     3361 atctgccggg tttccaccag ctgtctcttg atgaagccgg ccttatccag ttcgctcagg
     3421 ccgcctctct cggccttggt cagattgtcg aactttctct gggtaatcag cttggcgttc
     3481 agcagctgcc gccagtagtt cttcatcttc ttcacgacct cttcggaggg cacgttgtcg
     3541 ctcttgcccc ggttcttgtc gcttctggtc agcaccttgt tgtcgatgga gtcgtccttc
     3601 agaaagctct gaggcacgat atggtccaca tcgtagtcgg acagccggtt gatgtccagt
     3661 tcctggtcca cgtacatatc ccgcccattc tgcaggtagt acaggtacag cttctcgttc
     3721 tgcagctggg tgttttccac ggggtgttct ttcaggatct ggctgcccag ctctttgatg
     3781 ccctcttcga tccgcttcat tctctcgcgg ctgttcttct gtcccttctg ggtggtctgg
     3841 ttctctctgg ccatttcgat cacgatgttc tcgggcttgt gccggcccat cactttcacg
     3901 agctcgtcca ccaccttcac tgtctgcagg atgcccttct taatggcggg gctgccggcc
     3961 agattggcaa tgtgctcgtg caggctatcg ccctggccgg acacctgggc tttctggatg
     4021 tcctctttaa aggtcaggct gtcgtcgtgg atcagctgca tgaagtttct gttggcgaag
     4081 ccgtcggact tcaggaaatc caggattgtc ttgccggact gcttgtcccg gatgccgttg
     4141 atcagcttcc ggctcagcct gccccagccg gtgtatctcc gccgcttcag ctgcttcatc
     4201 actttgtcgt cgaacaggtg ggcataggtt ttcagccgtt cctcgatcat ctctctgtcc
     4261 tcaaacagtg tcagggtcag cacgatatct tccagaatgt cctcgttttc ctcattgtcc
     4321 aggaagtcct tgtccttgat aattttcagc agatcgtggt atgtgcccag ggaggcgttg
     4381 aaccgatctt ccacgccgga gatttccacg gagtcgaagc actcgatttt cttgaagtag
     4441 tcctctttca gctgcttcac ggtcactttc cggttggtct tgaacagcag gtccacgatg
     4501 gcctttttct gctcgccgct caggaaggcg ggctttctca ttccctcggt cacgtatttc
     4561 actttggtca gctcgttata cacggtgaag tactcgtaca gcaggctgtg cttgggcagc
     4621 accttctcgt tgggcaggtt cttatcgaag ttggtcatcc gctcgatgaa gctctgggcg
     4681 gaagcgccct tgtccaccac ttcctcgaag ttccaggggg tgatggtttc ctcgctcttt
     4741 ctggtcatcc aggcgaatct gctgtttccc ctggccagag ggcccacgta gtaggggatg
     4801 cggaaggtca ggatcttctc gatcttttcc cggttgtcct tcaggaatgg gtaaaaatct
     4861 tcctgccgcc gcagaatggc gtgcagctct cccaggtgga tctggtgggg gatgctgccg
     4921 ttgtcgaagg tccgctgctt ccgcagcagg tcctctctgt tcagcttcac gagcagttcc
     4981 tcggtgccgt ccatcttttc caggatgggc ttgatgaact tgtagaactc ttcctggctg
     5041 gctccgccgt caatgtagcc ggcgtagccg ttcttgctct ggtcgaagaa aatctctttg
     5101 tacttctcag gcagctgctg ccgcacgaga gctttcagca gggtcaggtc ctggtggtgc
     5161 tcgtcgtatc tcttgatcat agaggcgctc aggggggcct tggtgatctc ggtgttcact
     5221 ctcaggatgt cgctcagcag gatggcgtcg gacaggttct tggcggccag aaacaggtcg
     5281 gcgtactggt cgccgatctg ggccagcagg ttgtccaggt cgtcgtcgta ggtgtccttg
     5341 ctcagctgca gtttggcatc ctcggccagg tcgaagttgc tcttgaagtt gggggtcagg
     5401 cccaggctca gggcaatcag gtttccgaac aggccattct tcttctcgcc gggcagctgg
     5461 gcgatcagat tttccagccg tctgctcttg ctcagtctgg cagacaggat ggccttggcg
     5521 tccacgccgc tggcgttgat ggggttttcc tcgaacagct ggttgtaggt ctgcaccagc
     5581 tggatgaaca gcttgtccac gtcgctgttg tcggggttca ggtcgccctc gatcaggaag
     5641 tggccccgga acttgatcat gtgggccagg gccagataga tcagccgcag gtcggccttg
     5701 tcggtgctgt ccaccagttt ctttctcagg tggtagatgg tggggtactt ctcgtggtag
     5761 gccacctcgt ccacgatgtt gccgaagatg gggtgccgct cgtgcttctt atcctcttcc
     5821 accaggaagg actcttccag tctgtggaag aagctgtcgt ccaccttggc catctcgttg
     5881 ctgaagatct cttgcagata gcagatccgg ttcttccgtc tggtgtatct tcttctggcg
     5941 gttctcttca gccgggtggc ctcggctgtt tcgccgctgt cgaacagcag ggctccgatc
     6001 aggttcttct tgatgctgtg ccggtcggtg ttgcccagca ccttgaattt cttgctgggc
     6061 accttgtact cgtcggtgat cacggcccag cccacagagt tggtgccgat gtccaggccg
     6121 atgctgtact tcttgtcggc tgctgggact ccgtggatac cgaccttccg cttcttcttt
     6181 ggggccatgg tggcggcctc agaggccaac agcggccgcg atatcctcga ggaattcttt
     6241 gccaaaatga tgagacagca caataaccag cacgttgccc aggagctgta ggaaaaagaa
     6301 gaaggcatga acatggttag cagaggctct agagccgccg gtcacacgcc agaagccgaa
     6361 ccccgccctg ccccgtcccc cccgaaggca gccgtccccc cgcggacagc cccgaggctg
     6421 gagagggaga aggggacggc ggcgcggcga cgcacgaagg ccctccccgc ccatttcctt
     6481 cctgccggcg ccgcaccgct tcgccccgcg cccgctagag ggggtgcggc ggcgcctccc
     6541 agatttcggc tccgcacaga tttgggacaa aggaagtccc tgcgccctct cgcacgatta
     6601 ccataaaagg caatggctgc ggctcgccgc gcctcgacag ccgccggcgc tccgggggcc
     6661 gccgcgcccc ycccccgmgc cccccccggc ccgmggcggc cccgccccgc ccggcacccc
     6721 cmcctgccgc caccccccgc ccggcacggc gagccccgcg ccacgccccg tacggagccc
     6781 cgcacccgaa gccgggccgt gctcagcaac tcggggaggg gggtgcaggg ggggttgcag
     6841 cccgaccgac gcgcccacac cccctgctca cccccccacg cacacacccc gcacgcagcc
     6901 tttgttcccc tcgcagcccc ccccgcaccg cggggcaccg cccccggccg cgctcccctc
     6961 gcgcacactg cggagcgcac aaagccccgc gccgcgcccg cagcgctcac agccgccggg
     7021 cagcgcggag ccgcacgcgg cgctccccac gcacacacac acgcacgcac cccccgagcc
     7081 gctccccccg cacaaagggc cctcccggag cccctcaagg ctttcacgca gccacagaaa
     7141 agaaacaagc cgtcattaaa ccaagcgcta attacagccc ggaggagaag ggccgtcccg
     7201 cccgctcacc tgtgggagta acgcggtcag tcagagccgg ggcgggcggc gcgaggcggc
     7261 ggcggagcgg ggcacggggc gaaggcagcg cgcagcgact cccgcccgcc gcgcgcttcg
     7321 ctttttatag ggccgccgcc gccgccgcct cgccataaaa ggaaactttc ggagcgcgcc
     7381 gctctgattg gctgccgccs cacctctccs cctccccccc ccccccccct cccccccccc
     7441 ccccccccct kgcgcgcccc cccccccccc ccgcccccat cgctgcacaa aataattaaa
     7501 aaataaataa atacaaaatt gggggtgggg agggggggga gatggggaga gtgaagcaga
     7561 acgtggggct cacctcgacc atggtaatag cgatgactaa tacgtagatg tactgccaag
     7621 taggaaagtc ccataaggtc atgtactggg cataatgcca ggcgggccat ttaccgtcat
     7681 tgacgtcaat agggggcgta cttggcatat gatacacttg atgtactgcc aagtgggcag
     7741 tttaccgtaa atactccacc cattgacgtc aatggaaagt ccctattggc gttactatgg
     7801 gaacatacgt cattattgac gtcaatgggc gggggtcgtt gggcggtcag ccaggcgggc
     7861 catttaccgt aagttatgta acgcggaact ccatatatgg gctatgaact aatgaccccg
     7921 taattgatta ctattaataa ctagtcaata atcaatgtcg actcgtattc ccgggctgcc
     7981 aagcagaaaa gccaccgctg aggagactcc ggtcactgtc ctcgccccgc ctcccccttc
     8041 cctccccttg gggaccaccg ggcgccacgc cgcgaacggt aagtgccgcg gtcgtcggcg
     8101 cctccgccct ccccctaggg ccccaattcc cagcgggcgc ggcgccggcc cctccccccg
     8161 ccgcgcgcgc gcccgctgcc ccgcccttcg tggccgcccg gcgtgggcgg tgccacccct
     8221 ccccccggcg gccccgcgcg cagctcccgg ctccctcccc cttcggatgt ggcttgagct
     8281 gtaggcgcgg agggccggag acgctgcaga cccgcgaccc ggagcagctc ggaggcggtg
     8341 aagtcggtgg ctttccttct ctctagctct cgctcgctgg tggtgcttca gatgccacac
     8401 gcgtagctta taataccggt tcaattgccg acccctcccc ccaacttctc ggggactgtg
     8461 ggcgatgtgc gctctgccca ctgacgggca ccggagcctc taggcgagac cctgtctcac
     8521 aaaataaagt aagcccggac tgagtgcgga aaggcgggcc tggcgggtct ggtctcccca
     8581 tgcgggccac cagaggccct gcagccttca gtcgcttgaa ggggtaatgg cgcttccact
     8641 cacaaacatg gcggacagag cgtgtgaacg agatgaacag cccctcaaaa atatggccgc
     8701 cgaggctgga cggccgtgcc ccagcagcac cgcctccgcg ccccacgtga tctctcgccg
     8761 ggcacagcgc tgaccgcgga ggtccaaccg gaagaatgtc cggattggac attcggaaga
     8821 gggcccgcct tccctgggga atctctgcgc acgcgcagaa cgcttcgacc aatgaaaaca
     8881 caggaagccg tccgcgcaac cgcgttgcgt cacttctgcc gcccctgttt caaggtatat
     8941 agccgtagac ggaacttcgc ctttctctcg gccttagcgc catttttttg ggtgagtgtt
     9001 ttttggttcc tgcgttggga ttccgtgtac aatccataga catctgacct cggcacttag
     9061 catcatcaca gcaaactaac tgtagccttt ctctctttcc ctgtagaaac ctctgcacct
     9121 gaggccacca tgttgagcaa gggcgaggag gtcatcaaag agttcatgcg cttcaaggtg
     9181 cgcatggagg gctccatgaa cggccatgag ttcgagatcg agggcgaggg cgagggccgc
     9241 ccctacgagg gcacccagac cgccaagctg aaggtgacca agggcggccc cctgcccttc
     9301 gcctgggaca tcctgtcccc ccagttcatg tacggctcca aggcgtacgt gaagcacccc
     9361 gccgacatcc ccgattacaa gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg
     9421 atgaacttcg aggacggcgg tctggtgacc gtgacccagg actcctccct gcaggacggc
     9481 acgctgatct acaaggtgaa gatgcgcggc accaacttcc cccccgacgg ccccgtaatg
     9541 cagaagaaga caatgggctg ggaggcctcc accgagcgcc tgtacccccg cgacggcgtg
     9601 ctgaagggcg agatccacca ggccctgaag ctgaaggacg gcggccacta cctggtggag
     9661 ttcaagacca tctacatggc caagaagccc gtgcaactgc ccggctacta ctacgtggac
     9721 accaagctgg acatcacctc ccacaacgag gactacacca tcgtggaaca gtacgagcgc
     9781 tccgagggcc gccaccacct gttcctgtac ggcatggacg agctgtacaa gggcagtgga
     9841 gctactaact tcagcctgct gaagcaggct ggtgacgtcg aggagaatcc tggccccatg
     9901 accgagtaca agcccacggt gcgcctcgcc acccgcgacg acgtccccag ggccgtacgc
     9961 accctcgccg ccgcgttcgc cgactacccc gccacgcgcc acaccgtcga tccggaccgc
    10021 cacatcgagc gggtcaccga gctgcaagaa ctcttcctca cgcgcgtcgg gctcgacatc
    10081 ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg tctggaccac gccggagagc
    10141 gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca tggccgagtt gagcggttcc
    10201 cggctggccg cgcagcaaca gatggaaggc ctcctggcgc cgcaccggcc caaggagccc
    10261 gcgtggttcc tggccaccgt cggcgtctcg cccgaccacc agggcaaggg tctgggcagc
    10321 gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg gggtgcccgc cttcctggag
    10381 acctccgcgc cccgcaacct ccccttctac gagcggctcg gcttcaccgt caccgccgac
    10441 gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc gcaagcccgg tgcctgattc
    10501 gaaggcctgt cgtgaattca ctcctcaggt gcaggctgcc tatcagaagg tggtggctgg
    10561 tgtggccaat gccctggctc acaaatacca ctgagatctt tttccctctg ccaaaaatta
    10621 tggggacatc atgaagcccc ttgagcatct gacttctggc taataaagga aatttatttt
    10681 cattgcaata gtgtgttgga attttttgtg tctctcactc ggaaggacat atgggagggc
    10741 aaatcattta aaacatcaga atgagtattt ggtttagagt ttggcaacat atgccatatg
    10801 ctggctgcca tgaactagct actcgggacc ccttaccgaa acatcgccgc attctgcaga
    10861 ggagtcgagt gtatgtaaac ttctgaccca ctgggaatgt gatgaaagaa ataaaagctg
    10921 aaatgaatca ttctctctac tattattctg atatttcaca ttcttaaaat aaagtggtga
    10981 tcctaactga cctaagacag ggaattttta ctaggattaa atgtcaggaa ttgtgaaaaa
    11041 gtgagtttaa atgtatttgg ctaaggtgta tgtaaacttc cgacttcaac tgtataggga
    11101 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
    11161 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
    11221 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
    11281 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
    11341 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
    11401 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
    11461 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
    11521 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
    11581 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
    11641 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
    11701 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
    11761 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
    11821 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
    11881 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
    11941 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
    12001 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
    12061 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
    12121 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
    12181 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
    12241 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
    12301 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
    12361 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
    12421 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
    12481 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
    12541 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
    12601 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
    12661 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
    12721 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
    12781 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
    12841 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
    12901 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
    12961 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
    13021 gtgccacctg atgcggtgtg aaataccgca cagatgcgta aggagaaaat accgcatcag
    13081 gaaattgtaa gcgttaatat tttgttaaaa tt
//
