LOCUS       sequence-331320-       12576 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  All-in-one doxycycline inducible lentiviral vector for expression of
            Cas9 in combination with turbo GFP using the P2A self-cleaving 
            peptide.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12576)
  AUTHORS   Niu Y, Ferreira Azevedo CA, Li X, Kamali E, Haagen Nielsen O, 
            Storgaard Sorensen C, Frodin M
  TITLE     Multiparametric and accurate functional analysis of genetic sequence
            variants using CRISPR-Select.
  JOURNAL   Nat Genet. 2022 Dec 5. doi: 10.1038/s41588-022-01224-7.
  PUBMED    36471068
REFERENCE   2  (bases 1 to 12576)
  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: "Nat Genet. 
            2022 Dec 5. doi: 10.1038/s41588-022-01224-7."
COMMENT     Sequence Label: pCW57-GFP-2A-Cas9
FEATURES             Location/Qualifiers
     source          1..12576
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        6..320
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    14..32
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    50..68
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    85..103
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    121..139
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    157..175
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    192..210
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    228..246
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     293..317
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      343..352
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             349..1044
                     /codon_start=1
                     /product="improved green fluorescent protein from 
                     Pontellina plumata (Evdokimov et al., 2006)"
                     /label=TurboGFP
                     /note="mammalian codon-optimized"
                     /translation="MESDESGLPAMEIECRITGTLNGVEFELVGGGEGTPEQGRMTNKM
                     KSTKGALTFSPYLLSHVMGYGFYHFGTYPSGYENPFLHAINNGGYTNTRIEKYEDGGVL
                     HVSFSYRYEAGRVIGDFKVMGTGFPEDSVIFTDKIIRSNATVEHLHPMGDNDLDGSFTR
                     TFSLRDGGYYSSVVDSHMHFKSAIHPSILQNGGPMFAFRRVEEDHSNTELGIVEYQHAF
                     KTPDADAGEE"
     CDS             1057..1113
                     /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             1114..1134
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1159..5259
                     /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             1159..5259
                     /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             5260..5307
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     promoter        5312..5822
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     5741..5759
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             5832..6428
                     /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(5832..5851)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     6328..6348
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             6429..6482
                     /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             join(6483..7100,7101..7106,7107..7142,7143..7145,7146..7181,
                     7182..7184,7185..7220,7221..7229)
                     /codon_start=1
                     /product="improved tetracycline-controlled transactivator"
                     /label=rtTA-Advanced
                     /note="In the Tet-On(R) system, rtTA-Advanced binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the presence of tetracycline or doxycycline."
                     /translation="MSRLDKSKVINGALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHFCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     primer_bind     complement(7269..7289)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(7687..7698)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             7903..8136
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    8208..8329
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8245..8264)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8299..8318
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      8369..8504
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        8504..8575
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(8576..8644,8645..9436)
                     /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(8794..8813)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      9607..10195
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     10096..10115
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     10349..10366
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    10483..10504
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(10519..10524,10525..10542,10543..10549)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    10557..10573
                     /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)."
     primer_bind     10562..10584
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     10581..10597
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     10581..10597
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     10616..10636
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        10618..10636
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        10664..10890
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             10891..11071
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    11118..11243
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    11736..11969
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             12154..12198
                     /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    12411..12528
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 atgtcgagtt tactccctat cagtgataga gaacgtatgt cgagtttact ccctatcagt
       61 gatagagaac gatgtcgagt ttactcccta tcagtgatag agaacgtatg tcgagtttac
      121 tccctatcag tgatagagaa cgtatgtcga gtttactccc tatcagtgat agagaacgta
      181 tgtcgagttt atccctatca gtgatagaga acgtatgtcg agtttactcc ctatcagtga
      241 tagagaacgt atgtcgaggt aggcgtgtac ggtgggaggc ctatataagc agagctcgtt
      301 tagtgaaccg tcagatcgcc tggagaattg gctagcgaat tcgccaccat ggagagcgac
      361 gagagcggcc tgcccgccat ggagatcgag tgccgcatca ccggcaccct gaacggcgtg
      421 gagttcgagc tggtgggcgg cggagagggc acccccgagc agggccgcat gaccaacaag
      481 atgaagagca ccaaaggcgc cctgaccttc agcccctacc tgctgagcca cgtgatgggc
      541 tacggcttct accacttcgg cacctacccc agcggctacg agaacccctt cctgcacgcc
      601 atcaacaacg gcggctacac caacacccgc atcgagaagt acgaggacgg cggcgtgctg
      661 cacgtgagct tcagctaccg ctacgaggcc ggccgcgtga tcggcgactt caaggtgatg
      721 ggcaccggct tccccgagga cagcgtgatc ttcaccgaca agatcatccg cagcaacgcc
      781 accgtggagc acctgcaccc catgggcgat aacgatctgg atggcagctt cacccgcacc
      841 ttcagcctgc gcgacggcgg ctactacagc tccgtggtgg acagccacat gcacttcaag
      901 agcgccatcc accccagcat cctgcagaac gggggcccca tgttcgcctt ccgccgcgtg
      961 gaggaggatc acagcaacac cgagctgggc atcgtggagt accagcacgc cttcaagacc
     1021 ccggatgcag atgccggtga agaaaccggt tccggagcca cgaacttctc tctgttaaag
     1081 caagcaggag acgtggaaga aaaccccggt cctccaaaga agaagcggaa ggtcggtatc
     1141 cacggagtcc cagcagccga caagaagtac agcatcggcc tggacatcgg caccaactct
     1201 gtgggctggg ccgtgatcac cgacgagtac aaggtgccca gcaagaaatt caaggtgctg
     1261 ggcaacaccg accggcacag catcaagaag aacctgatcg gagccctgct gttcgacagc
     1321 ggcgaaacag ccgaggccac ccggctgaag agaaccgcca gaagaagata caccagacgg
     1381 aagaaccgga tctgctatct gcaagagatc ttcagcaacg agatggccaa ggtggacgac
     1441 agcttcttcc acagactgga agagtccttc ctggtggaag aggataagaa gcacgagcgg
     1501 caccccatct tcggcaacat cgtggacgag gtggcctacc acgagaagta ccccaccatc
     1561 taccacctga gaaagaaact ggtggacagc accgacaagg ccgacctgcg gctgatctat
     1621 ctggccctgg cccacatgat caagttccgg ggccacttcc tgatcgaggg cgacctgaac
     1681 cccgacaaca gcgacgtgga caagctgttc atccagctgg tgcagaccta caaccagctg
     1741 ttcgaggaaa accccatcaa cgccagcggc gtggacgcca aggccatcct gtctgccaga
     1801 ctgagcaaga gcagacggct ggaaaatctg atcgcccagc tgcccggcga gaagaagaat
     1861 ggcctgttcg gaaacctgat tgccctgagc ctgggcctga cccccaactt caagagcaac
     1921 ttcgacctgg ccgaggatgc caaactgcag ctgagcaagg acacctacga cgacgacctg
     1981 gacaacctgc tggcccagat cggcgaccag tacgccgacc tgtttctggc cgccaagaac
     2041 ctgtccgacg ccatcctgct gagcgacatc ctgagagtga acaccgagat caccaaggcc
     2101 cccctgagcg cctctatgat caagagatac gacgagcacc accaggacct gaccctgctg
     2161 aaagctctcg tgcggcagca gctgcctgag aagtacaaag agattttctt cgaccagagc
     2221 aagaacggct acgccggcta cattgacggc ggagccagcc aggaagagtt ctacaagttc
     2281 atcaagccca tcctggaaaa gatggacggc accgaggaac tgctcgtgaa gctgaacaga
     2341 gaggacctgc tgcggaagca gcggaccttc gacaacggca gcatccccca ccagatccac
     2401 ctgggagagc tgcacgccat tctgcggcgg caggaagatt tttacccatt cctgaaggac
     2461 aaccgggaaa agatcgagaa gatcctgacc ttccgcatcc cctactacgt gggccctctg
     2521 gccaggggaa acagcagatt cgcctggatg accagaaaga gcgaggaaac catcaccccc
     2581 tggaacttcg aggaagtggt ggacaagggc gcttccgccc agagcttcat cgagcggatg
     2641 accaacttcg ataagaacct gcccaacgag aaggtgctgc ccaagcacag cctgctgtac
     2701 gagtacttca ccgtgtataa cgagctgacc aaagtgaaat acgtgaccga gggaatgaga
     2761 aagcccgcct tcctgagcgg cgagcagaaa aaggccatcg tggacctgct gttcaagacc
     2821 aaccggaaag tgaccgtgaa gcagctgaaa gaggactact tcaagaaaat cgagtgcttc
     2881 gactccgtgg aaatctccgg cgtggaagat cggttcaacg cctccctggg cacataccac
     2941 gatctgctga aaattatcaa ggacaaggac ttcctggaca atgaggaaaa cgaggacatt
     3001 ctggaagata tcgtgctgac cctgacactg tttgaggaca gagagatgat cgaggaacgg
     3061 ctgaaaacct atgcccacct gttcgacgac aaagtgatga agcagctgaa gcggcggaga
     3121 tacaccggct ggggcaggct gagccggaag ctgatcaacg gcatccggga caagcagtcc
     3181 ggcaagacaa tcctggattt cctgaagtcc gacggcttcg ccaacagaaa cttcatgcag
     3241 ctgatccacg acgacagcct gacctttaaa gaggacatcc agaaagccca ggtgtccggc
     3301 cagggcgata gcctgcacga gcacattgcc aatctggccg gcagccccgc cattaagaag
     3361 ggcatcctgc agacagtgaa ggtggtggac gagctcgtga aagtgatggg ccggcacaag
     3421 cccgagaaca tcgtgatcga aatggccaga gagaaccaga ccacccagaa gggacagaag
     3481 aacagccgcg agagaatgaa gcggatcgaa gagggcatca aagagctggg cagccagatc
     3541 ctgaaagaac accccgtgga aaacacccag ctgcagaacg agaagctgta cctgtactac
     3601 ctgcagaatg ggcgggatat gtacgtggac caggaactgg acatcaaccg gctgtccgac
     3661 tacgatgtgg accatatcgt gcctcagagc tttctgaagg acgactccat cgacaacaag
     3721 gtgctgacca gaagcgacaa gaaccggggc aagagcgaca acgtgccctc cgaagaggtc
     3781 gtgaagaaga tgaagaacta ctggcggcag ctgctgaacg ccaagctgat tacccagaga
     3841 aagttcgaca atctgaccaa ggccgagaga ggcggcctga gcgaactgga taaggccggc
     3901 ttcatcaaga gacagctggt ggaaacccgg cagatcacaa agcacgtggc acagatcctg
     3961 gactcccgga tgaacactaa gtacgacgag aatgacaagc tgatccggga agtgaaagtg
     4021 atcaccctga agtccaagct ggtgtccgat ttccggaagg atttccagtt ttacaaagtg
     4081 cgcgagatca acaactacca ccacgcccac gacgcctacc tgaacgccgt cgtgggaacc
     4141 gccctgatca aaaagtaccc taagctggaa agcgagttcg tgtacggcga ctacaaggtg
     4201 tacgacgtgc ggaagatgat cgccaagagc gagcaggaaa tcggcaaggc taccgccaag
     4261 tacttcttct acagcaacat catgaacttt ttcaagaccg agattaccct ggccaacggc
     4321 gagatccgga agcggcctct gatcgagaca aacggcgaaa ccggggagat cgtgtgggat
     4381 aagggccggg attttgccac cgtgcggaaa gtgctgagca tgccccaagt gaatatcgtg
     4441 aaaaagaccg aggtgcagac aggcggcttc agcaaagagt ctatcctgcc caagaggaac
     4501 agcgataagc tgatcgccag aaagaaggac tgggacccta agaagtacgg cggcttcgac
     4561 agccccaccg tggcctattc tgtgctggtg gtggccaaag tggaaaaggg caagtccaag
     4621 aaactgaaga gtgtgaaaga gctgctgggg atcaccatca tggaaagaag cagcttcgag
     4681 aagaatccca tcgactttct ggaagccaag ggctacaaag aagtgaaaaa ggacctgatc
     4741 atcaagctgc ctaagtactc cctgttcgag ctggaaaacg gccggaagag aatgctggcc
     4801 tctgccggcg aactgcagaa gggaaacgaa ctggccctgc cctccaaata tgtgaacttc
     4861 ctgtacctgg ccagccacta tgagaagctg aagggctccc ccgaggataa tgagcagaaa
     4921 cagctgtttg tggaacagca caagcactac ctggacgaga tcatcgagca gatcagcgag
     4981 ttctccaaga gagtgatcct ggccgacgct aatctggaca aagtgctgtc cgcctacaac
     5041 aagcaccggg ataagcccat cagagagcag gccgagaata tcatccacct gtttaccctg
     5101 accaatctgg gagcccctgc cgccttcaag tactttgaca ccaccatcga ccggaagagg
     5161 tacaccagca ccaaagaggt gctggacgcc accctgatcc accagagcat caccggcctg
     5221 tacgagacac ggatcgacct gtctcagctg ggaggcgaca aaaggccggc ggccacgaaa
     5281 aaggccggcc aggcaaaaaa gaaaaagtaa cggggttggg gttgcgcctt ttccaaggca
     5341 gccctgggtt tgcgcaggga cgcggctgct ctgggcgtgg ttccgggaaa cgcagcggcg
     5401 ccgaccctgg gtctcgcaca ttcttcacgt ccgttcgcag cgtcacccgg atcttcgccg
     5461 ctacccttgt gggccccccg gcgacgcttc ctgctccgcc cctaagtcgg gaaggttcct
     5521 tgcggttcgc ggcgtgccgg acgtgacaaa cggaagccgc acgtctcact agtaccctcg
     5581 cagacggaca gcgccaggga gcaatggcag cgcgccgacc gcgatgggct gtggccaata
     5641 gcggctgctc agcagggcgc gccgagagca gcggccggga aggggcggtg cgggaggcgg
     5701 ggtgtggggc ggtagtgtgg gccctgttcc tgcccgcgcg gtgttccgca ttctgcaagc
     5761 ctccggagcg cacgtcggca gtcggctccc tcgttgaccg aatcaccgac ctctctcccc
     5821 agcaattcac catgaccgag tacaagccca cggtgcgcct cgccacccgc gacgacgtcc
     5881 ccagggccgt acgcaccctc gccgccgcgt tcgccgacta ccccgccacg cgccacaccg
     5941 tcgatccgga ccgccacatc gagcgggtca ccgagctgca agaactcttc ctcacgcgcg
     6001 tcgggctcga catcggcaag gtgtgggtcg cggacgacgg cgccgcggtg gcggtctgga
     6061 ccacgccgga gagcgtcgaa gcgggggcgg tgttcgccga gatcggcccg cgcatggccg
     6121 agttgagcgg ttcccggctg gccgcgcagc aacagatgga aggcctcctg gcgccgcacc
     6181 ggcccaagga gcccgcgtgg ttcctggcca ccgtcggcgt ctcgcccgac caccagggca
     6241 agggtctggg cagcgccgtc gtgctccccg gagtggaggc ggccgagcgc gccggggtgc
     6301 ccgccttcct ggagacctcc gcgccccgca acctcccctt ctacgagcgg ctcggcttca
     6361 ccgtcaccgc cgacgtcgag gtgcccgaag gaccgcgcac ctggtgcatg acccgcaagc
     6421 ccggtgccga aggtagaggt tctctcctca cttgtggtga tgttgaagaa aaccctggtc
     6481 caatgtctag actggacaag agcaaagtca taaacggagc tctggaatta ctcaatggtg
     6541 tcggtatcga aggcctgacg acaaggaaac tcgctcaaaa gctgggagtt gagcagccta
     6601 ccctgtactg gcacgtgaag aacaagcggg ccctgctcga tgccctgcca atcgagatgc
     6661 tggacaggca tcatacccac ttctgccccc tggaaggcga gtcatggcaa gactttctgc
     6721 ggaacaacgc caagtcatac cgctgtgctc tcctctcaca tcgcgacggg gctaaagtgc
     6781 atctcggcac ccgcccaaca gagaaacagt acgaaaccct ggaaaatcag ctcgcgttcc
     6841 tgtgtcagca aggcttctcc ctggagaacg cactgtacgc tctgtccgcc gtgggccact
     6901 ttacactggg ctgcgtattg gaggaacagg agcatcaagt agcaaaagag gaaagagaga
     6961 cacctaccac cgattctatg cccccacttc tgagacaagc aattgagctg ttcgaccggc
     7021 agggagccga acctgccttc cttttcggcc tggaactaat catatgtggc ctggagaaac
     7081 agctaaagtg cgaaagcggc gggccgaccg acgcccttga cgattttgac ttagacatgc
     7141 tcccagccga tgcccttgac gactttgacc ttgatatgct gcctgctgac gctcttgacg
     7201 attttgacct tgacatgctc cccgggtaac taagtaagga tcgatccaag atatcgtatt
     7261 cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat
     7321 gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct
     7381 ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct
     7441 gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc
     7501 gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg
     7561 acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa gctgacgtcc
     7621 tttccatggc tgctcgcctg tgttgccacc tggattctgc gcgggacgtc cttctgctac
     7681 gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg
     7741 cctcttccgc gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc
     7801 ccgcctgttt cgcctcggcg tccggactag aggtaccttt aagaccaatg acttacaagg
     7861 cagctgtaga tcttagccac tttttaaaag aaaagggggg actggaaggg ctaattcact
     7921 cccaacgaag acaagatctg ctttttgctt gtactgggtc tctctggtta gaccagatct
     7981 gagcctggga gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc
     8041 cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc
     8101 tcagaccctt ttagtcagtg tggaaaatct ctagcagtag tagttcatgt catcttatta
     8161 ttcagtattt ataacttgca aagaaatgaa tatcagagag tgagaggaac ttgtttattg
     8221 cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt
     8281 tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctggc
     8341 tctagctatc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     8401 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc
     8461 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctttcaa atatgtatcc
     8521 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
     8581 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     8641 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     8701 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     8761 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     8821 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     8881 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     8941 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     9001 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     9061 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     9121 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     9181 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     9241 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     9301 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     9361 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     9421 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     9481 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     9541 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     9601 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     9661 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     9721 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     9781 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     9841 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     9901 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     9961 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
    10021 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
    10081 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
    10141 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
    10201 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
    10261 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
    10321 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg
    10381 cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga
    10441 caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga gttagctcac
    10501 tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt
    10561 gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca agcgcgcaat
    10621 taaccctcac taaagggaac aaaagctgga gctgcaagct taatgtagtc ttatgcaata
    10681 ctcttgtagt cttgcaacat ggtaacgatg agttagcaac atgccttaca aggagagaaa
    10741 aagcaccgtg catgccgatt ggtggaagta aggtggtacg atcgtgcctt attaggaagg
    10801 caacagacgg gtctgacatg gattggacga accactgaat tgccgcattg cagagatatt
    10861 gtatttaagt gcctagctcg atacataaac gggtctctct ggttagacca gatctgagcc
    10921 tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag cttgccttga
    10981 gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag atccctcaga
    11041 cccttttagt cagtgtggaa aatctctagc agtggcgccc gaacagggac ttgaaagcga
    11101 aagggaaacc agaggagctc tctcgacgca ggactcggct tgctgaagcg cgcacggcaa
    11161 gaggcgaggg gcggcgactg gtgagtacgc caaaaatttt gactagcgga ggctagaagg
    11221 agagagatgg gtgcgagagc gtcagtatta agcgggggag aattagatcg cgatgggaaa
    11281 aaattcggtt aaggccaggg ggaaagaaaa aatataaatt aaaacatata gtatgggcaa
    11341 gcagggagct agaacgattc gcagttaatc ctggcctgtt agaaacatca gaaggctgta
    11401 gacaaatact gggacagcta caaccatccc ttcagacagg atcagaagaa cttagatcat
    11461 tatataatac agtagcaacc ctctattgtg tgcatcaaag gatagagata aaagacacca
    11521 aggaagcttt agacaagata gaggaagagc aaaacaaaag taagaccacc gcacagcaag
    11581 cggccgctga tcttcagacc tggaggagga gatatgaggg acaattggag aagtgaatta
    11641 tataaatata aagtagtaaa aattgaacca ttaggagtag cacccaccaa ggcaaagaga
    11701 agagtggtgc agagagaaaa aagagcagtg ggaataggag ctttgttcct tgggttcttg
    11761 ggagcagcag gaagcactat gggcgcagcg tcaatgacgc tgacggtaca ggccagacaa
    11821 ttattgtctg gtatagtgca gcagcagaac aatttgctga gggctattga ggcgcaacag
    11881 catctgttgc aactcacagt ctggggcatc aagcagctcc aggcaagaat cctggctgtg
    11941 gaaagatacc taaaggatca acagctcctg gggatttggg gttgctctgg aaaactcatt
    12001 tgcaccactg ctgtgccttg gaatgctagt tggagtaata aatctctgga acagatttgg
    12061 aatcacacga cctggatgga gtgggacaga gaaattaaca attacacaag cttaatacac
    12121 tccttaattg aagaatcgca aaaccagcaa gaaaagaatg aacaagaatt attggaatta
    12181 gataaatggg caagtttgtg gaattggttt aacataacaa attggctgtg gtatataaaa
    12241 ttattcataa tgatagtagg aggcttggta ggtttaagaa tagtttttgc tgtactttct
    12301 atagtgaata gagttaggca gggatattca ccattatcgt ttcagaccca cctcccaacc
    12361 ccgaggggac aattctcgac ctcgagacaa atggcagtat tcatccacaa ttttaaaaga
    12421 aaagggggga ttggggggta cagtgcaggg gaaagaatag tagacataat agcaacagac
    12481 atacaaacta aagaattaca aaaacaaatt acaaaaattc aaaattttcg ggtttattac
    12541 agggacagca gagatccact ttggccgcga atcgat
//
