LOCUS       sequence-361330-        9347 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cas9 from S.pyogenes with 2A-Puro, and the 2A-sgRNA to targeting 
            exon 2 of human HLA-G gene.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9347)
  AUTHORS   Palma MB, Tronik-Le Roux D, Amin G, Castaneda S, Mobbs AM, Scarafia 
            MA, La Greca A, Daouya M, Poras I, Inda AM, Moro LN, Carosella ED, 
            Garcia MN, Miriuka SG
  TITLE     HLA-G gene editing in tumor cell lines as a novel alternative in 
            cancer immunotherapy.
  JOURNAL   Sci Rep. 2021 Nov 12;11(1):22158. doi: 10.1038/s41598-021-01572-0.
  PUBMED    34773056
REFERENCE   2  (bases 1 to 9347)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Sci Rep."; 
            date: "2021-11-12"; volume: "11(1):22158. doi"; pages: " 
            10.1038/s41598-021-01572-0"
COMMENT     Sequence Label: pSpCas9-Puro HLAG-2A-sgRNA
FEATURES             Location/Qualifiers
     source          1..9347
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        117..365
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     117..137
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     288..307
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        387..462
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        559..844
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     intron          1294..1521
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      1533..1542
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1542..1607
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1614..1634
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1659..5759
                     /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             1659..5759
                     /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             5760..5807
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             5823..5876
                     /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             5877..6476
                     /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(5877..5896)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6373..6393
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(6504..6521)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6510..6717
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   6726..6866
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   6726..6855
                     /label=AAV2 ITR
     rep_origin      6941..7396
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7028..7047)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7238..7259
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(7413..7432)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7532..7554
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7592..7610)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7678..7782
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7783..7851,7852..8643)
                     /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     complement(8001..8020)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(8814..9347,1..55)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9303..9322
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
       61 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgtgagg
      121 gcctatttcc catgattcct tcatatttgc atatacgata caaggctgtt agagagataa
      181 ttggaattaa tttgactgta aacacaaaga tattagtaca aaatacgtga cgtagaaagt
      241 aataatttct tgggtagttt gcagttttaa aattatgttt taaaatggac tatcatatgc
      301 ttaccgtaac ttgaaagtat ttcgatttct tggctttata tatcttgtgg aaaggacgaa
      361 acaccgtggg gagaatgagt ccgggtgttt tagagctaga aatagcaagt taaaataagg
      421 ctagtccgtt atcaacttga aaaagtggca ccgagtcggt gcttttttgt tttagagcta
      481 gaaatagcaa gttaaaataa ggctagtccg tttttagcgc gtgcgccaat tctgcagaca
      541 aatggctcta gaggtacccg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
      601 caacgacccc cgcccattga cgtcaatagt aacgccaata gggactttcc attgacgtca
      661 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      721 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt gtgcccagta
      781 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      841 catggtcgag gtgagcccca cgttctgctt cactctcccc atctcccccc cctccccacc
      901 cccaattttg tatttattta ttttttaatt attttgtgca gcgatggggg cggggggggg
      961 gggggggsgs ssnccngggg gggggggggg gggggggggg gggggggggg gggggggggg
     1021 gggggggggg gggggggggg gggggggggg gggggggggg gggggggggg gggggggggg
     1081 gggggggggg gggggggggg gggggggggg gggggggggg gggggggggg gggggggcgc
     1141 gcgccrggsg gggcggggcg gggcgagggg cggggcgggg cgaggcggag aggtgcggcg
     1201 gcagccaatc agagcggcgc gctccgaaag tttcctttta tggcgaggcg gcggcggcgg
     1261 cggccctata aaaagcgaag cgcgcggcgg gcgggagtcg ctgcgcgctg ccttcgcccc
     1321 gtgccccgct ccgccgccgc ctcgcgccgc ccgccccggc tctgactgac cgcgttactc
     1381 ccacaggtga gcgggcggga cggcccttct cctccgggct gtaattagct gagcaagagg
     1441 taagggttta agggatggtt ggttggtggg gtattaatgt ttaattacct ggagcacctg
     1501 cctgaaatca ctttttttca ggttggaccg gtgccaccat ggactataag gaccacgacg
     1561 gagactacaa ggatcatgat attgattaca aagacgatga cgataagatg gccccaaaga
     1621 agaagcggaa ggtcggtatc cacggagtcc cagcagccga caagaagtac agcatcggcc
     1681 tggacatcgg caccaactct gtgggctggg ccgtgatcac cgacgagtac aaggtgccca
     1741 gcaagaaatt caaggtgctg ggcaacaccg accggcacag catcaagaag aacctgatcg
     1801 gagccctgct gttcgacagc ggcgaaacag ccgaggccac ccggctgaag agaaccgcca
     1861 gaagaagata caccagacgg aagaaccgga tctgctatct gcaagagatc ttcagcaacg
     1921 agatggccaa ggtggacgac agcttcttcc acagactgga agagtccttc ctggtggaag
     1981 aggataagaa gcacgagcgg caccccatct tcggcaacat cgtggacgag gtggcctacc
     2041 acgagaagta ccccaccatc taccacctga gaaagaaact ggtggacagc accgacaagg
     2101 ccgacctgcg gctgatctat ctggccctgg cccacatgat caagttccgg ggccacttcc
     2161 tgatcgaggg cgacctgaac cccgacaaca gcgacgtgga caagctgttc atccagctgg
     2221 tgcagaccta caaccagctg ttcgaggaaa accccatcaa cgccagcggc gtggacgcca
     2281 aggccatcct gtctgccaga ctgagcaaga gcagacggct ggaaaatctg atcgcccagc
     2341 tgcccggcga gaagaagaat ggcctgttcg gaaacctgat tgccctgagc ctgggcctga
     2401 cccccaactt caagagcaac ttcgacctgg ccgaggatgc caaactgcag ctgagcaagg
     2461 acacctacga cgacgacctg gacaacctgc tggcccagat cggcgaccag tacgccgacc
     2521 tgtttctggc cgccaagaac ctgtccgacg ccatcctgct gagcgacatc ctgagagtga
     2581 acaccgagat caccaaggcc cccctgagcg cctctatgat caagagatac gacgagcacc
     2641 accaggacct gaccctgctg aaagctctcg tgcggcagca gctgcctgag aagtacaaag
     2701 agattttctt cgaccagagc aagaacggct acgccggcta cattgacggc ggagccagcc
     2761 aggaagagtt ctacaagttc atcaagccca tcctggaaaa gatggacggc accgaggaac
     2821 tgctcgtgaa gctgaacaga gaggacctgc tgcggaagca gcggaccttc gacaacggca
     2881 gcatccccca ccagatccac ctgggagagc tgcacgccat tctgcggcgg caggaagatt
     2941 tttacccatt cctgaaggac aaccgggaaa agatcgagaa gatcctgacc ttccgcatcc
     3001 cctactacgt gggccctctg gccaggggaa acagcagatt cgcctggatg accagaaaga
     3061 gcgaggaaac catcaccccc tggaacttcg aggaagtggt ggacaagggc gcttccgccc
     3121 agagcttcat cgagcggatg accaacttcg ataagaacct gcccaacgag aaggtgctgc
     3181 ccaagcacag cctgctgtac gagtacttca ccgtgtataa cgagctgacc aaagtgaaat
     3241 acgtgaccga gggaatgaga aagcccgcct tcctgagcgg cgagcagaaa aaggccatcg
     3301 tggacctgct gttcaagacc aaccggaaag tgaccgtgaa gcagctgaaa gaggactact
     3361 tcaagaaaat cgagtgcttc gactccgtgg aaatctccgg cgtggaagat cggttcaacg
     3421 cctccctggg cacataccac gatctgctga aaattatcaa ggacaaggac ttcctggaca
     3481 atgaggaaaa cgaggacatt ctggaagata tcgtgctgac cctgacactg tttgaggaca
     3541 gagagatgat cgaggaacgg ctgaaaacct atgcccacct gttcgacgac aaagtgatga
     3601 agcagctgaa gcggcggaga tacaccggct ggggcaggct gagccggaag ctgatcaacg
     3661 gcatccggga caagcagtcc ggcaagacaa tcctggattt cctgaagtcc gacggcttcg
     3721 ccaacagaaa cttcatgcag ctgatccacg acgacagcct gacctttaaa gaggacatcc
     3781 agaaagccca ggtgtccggc cagggcgata gcctgcacga gcacattgcc aatctggccg
     3841 gcagccccgc cattaagaag ggcatcctgc agacagtgaa ggtggtggac gagctcgtga
     3901 aagtgatggg ccggcacaag cccgagaaca tcgtgatcga aatggccaga gagaaccaga
     3961 ccacccagaa gggacagaag aacagccgcg agagaatgaa gcggatcgaa gagggcatca
     4021 aagagctggg cagccagatc ctgaaagaac accccgtgga aaacacccag ctgcagaacg
     4081 agaagctgta cctgtactac ctgcagaatg ggcgggatat gtacgtggac caggaactgg
     4141 acatcaaccg gctgtccgac tacgatgtgg accatatcgt gcctcagagc tttctgaagg
     4201 acgactccat cgacaacaag gtgctgacca gaagcgacaa gaaccggggc aagagcgaca
     4261 acgtgccctc cgaagaggtc gtgaagaaga tgaagaacta ctggcggcag ctgctgaacg
     4321 ccaagctgat tacccagaga aagttcgaca atctgaccaa ggccgagaga ggcggcctga
     4381 gcgaactgga taaggccggc ttcatcaaga gacagctggt ggaaacccgg cagatcacaa
     4441 agcacgtggc acagatcctg gactcccgga tgaacactaa gtacgacgag aatgacaagc
     4501 tgatccggga agtgaaagtg atcaccctga agtccaagct ggtgtccgat ttccggaagg
     4561 atttccagtt ttacaaagtg cgcgagatca acaactacca ccacgcccac gacgcctacc
     4621 tgaacgccgt cgtgggaacc gccctgatca aaaagtaccc taagctggaa agcgagttcg
     4681 tgtacggcga ctacaaggtg tacgacgtgc ggaagatgat cgccaagagc gagcaggaaa
     4741 tcggcaaggc taccgccaag tacttcttct acagcaacat catgaacttt ttcaagaccg
     4801 agattaccct ggccaacggc gagatccgga agcggcctct gatcgagaca aacggcgaaa
     4861 ccggggagat cgtgtgggat aagggccggg attttgccac cgtgcggaaa gtgctgagca
     4921 tgccccaagt gaatatcgtg aaaaagaccg aggtgcagac aggcggcttc agcaaagagt
     4981 ctatcctgcc caagaggaac agcgataagc tgatcgccag aaagaaggac tgggacccta
     5041 agaagtacgg cggcttcgac agccccaccg tggcctattc tgtgctggtg gtggccaaag
     5101 tggaaaaggg caagtccaag aaactgaaga gtgtgaaaga gctgctgggg atcaccatca
     5161 tggaaagaag cagcttcgag aagaatccca tcgactttct ggaagccaag ggctacaaag
     5221 aagtgaaaaa ggacctgatc atcaagctgc ctaagtactc cctgttcgag ctggaaaacg
     5281 gccggaagag aatgctggcc tctgccggcg aactgcagaa gggaaacgaa ctggccctgc
     5341 cctccaaata tgtgaacttc ctgtacctgg ccagccacta tgagaagctg aagggctccc
     5401 ccgaggataa tgagcagaaa cagctgtttg tggaacagca caagcactac ctggacgaga
     5461 tcatcgagca gatcagcgag ttctccaaga gagtgatcct ggccgacgct aatctggaca
     5521 aagtgctgtc cgcctacaac aagcaccggg ataagcccat cagagagcag gccgagaata
     5581 tcatccacct gtttaccctg accaatctgg gagcccctgc cgccttcaag tactttgaca
     5641 ccaccatcga ccggaagagg tacaccagca ccaaagaggt gctggacgcc accctgatcc
     5701 accagagcat caccggcctg tacgagacac ggatcgacct gtctcagctg ggaggcgaca
     5761 aaaggccggc ggccacgaaa aaggccggcc aggcaaaaaa gaaaaaggaa ttcggcagtg
     5821 gagagggcag aggaagtctg ctaacatgcg gtgacgtcga ggagaatcct ggcccaatga
     5881 ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga cgtccccagg gccgtacgca
     5941 ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca caccgtcgat ccggaccgcc
     6001 acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac gcgcgtcggg ctcgacatcg
     6061 gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt ctggaccacg ccggagagcg
     6121 tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat ggccgagttg agcggttccc
     6181 ggctggccgc gcagcaacag atggaaggcc tcctggcgcc gcaccggccc aaggagcccg
     6241 cgtggttcct ggccaccgtc ggagtctcgc ccgaccacca gggcaagggt ctgggcagcg
     6301 ccgtcgtgct ccccggagtg gaggcggccg agcgcgccgg ggtgcccgcc ttcctggaga
     6361 cctccgcgcc ccgcaacctc cccttctacg agcggctcgg cttcaccgtc accgccgacg
     6421 tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg caagcccggt gcctgagaat
     6481 tctaactaga gctcgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt
     6541 ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta
     6601 ataaaatgag gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg
     6661 ggtggggcag gacagcaagg gggaggattg ggaagagaat agcaggcatg ctggggagcg
     6721 gccgcaggaa cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac
     6781 tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag
     6841 cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg tattttctcc ttacgcatct
     6901 gtgcggtatt tcacaccgca tacgtcaaag caaccatagt acgcgccctg tagcggcgca
     6961 ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcctta
     7021 gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt
     7081 caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac
     7141 cccaaaaaac ttgatttggg tgatggttca cgtagtgggc catcgccctg atagacggtt
     7201 tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga
     7261 acaacactca actctatctc gggctattct tttgatttat aagggatttt gccgatttcg
     7321 gtctattggt taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata
     7381 ttaacgttta caattttatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
     7441 agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg
     7501 gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca
     7561 ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt
     7621 aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc
     7681 ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa
     7741 taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc
     7801 cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa
     7861 acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa
     7921 ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg
     7981 atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa
     8041 gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc
     8101 acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc
     8161 atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta
     8221 accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag
     8281 ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca
     8341 acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca acaattaata
     8401 gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc
     8461 tggtttattg ctgataaatc tggagccggt gagcgtggaa gccgcggtat cattgcagca
     8521 ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca
     8581 actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg
     8641 taactgtcag accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa
     8701 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     8761 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     8821 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     8881 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     8941 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     9001 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     9061 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     9121 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     9181 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     9241 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     9301 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacct
//
