LOCUS       sequence-418381-       12386 bp    DNA     circular SYN 15-APR-2025
DEFINITION  lentiviral vector expressing SpCas9 and tracrRNA for stable cell 
            line establishment.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12386)
  AUTHORS   Zhang H, Kelly K, Lee J, Echeverria D, Cooper D, Panwala R, Amrani 
            N, Chen Z, Gaston N, Wagh A, Newby GA, Xie J, Liu DR, Gao G, Wolfe 
            SA, Khvorova A, Watts JK, Sontheimer EJ
  TITLE     Self-delivering, chemically modified CRISPR RNAs for AAV co-delivery
            and genome editing in vivo.
  JOURNAL   Nucleic Acids Res. 2023 Nov 30:gkad1125. doi: 10.1093/nar/gkad1125.
  PUBMED    38033325
REFERENCE   2  (bases 1 to 12386)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nucleic 
            Acids Res. 2023 Nov 30:gkad1125. doi: 10.1093/nar/gkad1125."
COMMENT     Sequence Label: pEJS-HZ03-pLenti-U1a-mSpCas9-U6-tracrRNA-PuroR
FEATURES             Location/Qualifiers
     source          1..12386
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        3..229
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             230..410
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    457..582
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1075..1308
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1493..1537
                     /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"
     CDS             1973..1999
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2003..2023
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2033..6133
                     /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             2033..6133
                     /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             6137..6157
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(6177..6196)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    6242..6297
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     promoter        6331..6579
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     6331..6351
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     6502..6521
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_feature    6705..6822
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        6871..7381
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     7300..7318
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             7403..8002
                     /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(7403..7422)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     7899..7919
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     LTR             8130..8363
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    8435..8556
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8472..8491)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8526..8545
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      8596..8731
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(8751..8770)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(8752..8770)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(8780..8797)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(8780..8796)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8789..8811)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      8938..9393
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9025..9044)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9235..9256
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        9419..9523
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(9524..9592,9593..10384)
                     /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(9742..9761)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      10555..11143
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     11044..11063
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     11297..11314
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    11431..11452
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     mobile_element  11477..12244
                     /mobile_element_type="insertion sequence:IS1"
                     /label=IS1
                     /note="prokaryotic transposable element"
     promoter        join(12245..12249,12250..12267,12268..12274)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    12282..12298
                     /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     12287..12309
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     12306..12322
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     12306..12322
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     12341..12361
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        12343..12361
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 aatgtagtct tatgcaatac tcttgtagtc ttgcaacatg gtaacgatga gttagcaaca
       61 tgccttacaa ggagagaaaa agcaccgtgc atgccgattg gtggaagtaa ggtggtacga
      121 tcgtgcctta ttaggaaggc aacagacggg tctgacatgg attggacgaa ccactgaatt
      181 gccgcattgc agagatattg tatttaagtg cctagctcga tacataaacg ggtctctctg
      241 gttagaccag atctgagcct gggagctctc tggctaacta gggaacccac tgcttaagcc
      301 tcaataaagc ttgccttgag tgcttcaagt agtgtgtgcc cgtctgttgt gtgactctgg
      361 taactagaga tccctcagac ccttttagtc agtgtggaaa atctctagca gtggcgcccg
      421 aacagggact tgaaagcgaa agggaaacca gaggagctct ctcgacgcag gactcggctt
      481 gctgaagcgc gcacggcaag aggcgagggg cggcgactgg tgagtacgcc aaaaattttg
      541 actagcggag gctagaagga gagagatggg tgcgagagcg tcagtattaa gcgggggaga
      601 attagatcgc gatgggaaaa aattcggtta aggccagggg gaaagaaaaa atataaatta
      661 aaacatatag tatgggcaag cagggagcta gaacgattcg cagttaatcc tggcctgtta
      721 gaaacatcag aaggctgtag acaaatactg ggacagctac aaccatccct tcagacagga
      781 tcagaagaac ttagatcatt atataataca gtagcaaccc tctattgtgt gcatcaaagg
      841 atagagataa aagacaccaa ggaagcttta gacaagatag aggaagagca aaacaaaagt
      901 aagaccaccg cacagcaagc ggccgctgat cttcagacct ggaggaggag atatgaggga
      961 caattggaga agtgaattat ataaatataa agtagtaaaa attgaaccat taggagtagc
     1021 acccaccaag gcaaagagaa gagtggtgca gagagaaaaa agagcagtgg gaataggagc
     1081 tttgttcctt gggttcttgg gagcagcagg aagcactatg ggcgcagcgt caatgacgct
     1141 gacggtacag gccagacaat tattgtctgg tatagtgcag cagcagaaca atttgctgag
     1201 ggctattgag gcgcaacagc atctgttgca actcacagtc tggggcatca agcagctcca
     1261 ggcaagaatc ctggctgtgg aaagatacct aaaggatcaa cagctcctgg ggatttgggg
     1321 ttgctctgga aaactcattt gcaccactgc tgtgccttgg aatgctagtt ggagtaataa
     1381 atctctggaa cagatttgga atcacacgac ctggatggag tgggacagag aaattaacaa
     1441 ttacacaagc ttaatacact ccttaattga agaatcgcaa aaccagcaag aaaagaatga
     1501 acaagaatta ttggaattag ataaatgggc aagtttgtgg aattggttta acataacaaa
     1561 ttggctgtgg tatataaaat tattcataat gatagtagga ggcttggtag gtttaagaat
     1621 agtttttgct gtactttcta tagtgaatag agttaggcag ggatattcac cattatcgtt
     1681 tcagacccac ctcccaaccc cgagggatgg aggcggtact atgtagatga gaattcagga
     1741 gcaaactggg aaaagcaact gcttccaaat atttgtgatt tttacagtgt agttttggaa
     1801 aaactcttag cctaccaatt cttctaagtg ttttaaaatg tgggagccag tacacatgaa
     1861 gttatagagt gttttaatga ggcttaaata tttaccgtaa ctatgaaatg ctacgcatat
     1921 catgctgttc aggctccgtg gccacgcaac tcatactacc ggtgccacca tgtacccata
     1981 cgatgttcca gattacgctt cgccgaagaa aaagcgcaag gtcgaagcgt ccgacaagaa
     2041 gtacagcatc ggcctggaca tcggcaccaa ctctgtgggc tgggccgtga tcaccgacga
     2101 gtacaaggtg cccagcaaga aattcaaggt gctgggcaac accgaccggc acagcatcaa
     2161 gaagaacctg atcggagccc tgctgttcga cagcggcgaa acagccgagg ccacccggct
     2221 gaagagaacc gccagaagaa gatacaccag acggaagaac cggatctgct atctgcaaga
     2281 gatcttcagc aacgagatgg ccaaggtgga cgacagcttc ttccacagac tggaagagtc
     2341 cttcctggtg gaagaggata agaagcacga gcggcacccc atcttcggca acatcgtgga
     2401 cgaggtggcc taccacgaga agtaccccac catctaccac ctgagaaaga aactggtgga
     2461 cagcaccgac aaggccgacc tgcggctgat ctatctggcc ctggcccaca tgatcaagtt
     2521 ccggggccac ttcctgatcg agggcgacct gaaccccgac aacagcgacg tggacaagct
     2581 gttcatccag ctggtgcaga cctacaacca gctgttcgag gaaaacccca tcaacgccag
     2641 cggcgtggac gccaaggcca tcctgtctgc cagactgagc aagagcagac ggctggaaaa
     2701 tctgatcgcc cagctgcccg gcgagaagaa gaatggcctg ttcggaaacc tgattgccct
     2761 gagcctgggc ctgaccccca acttcaagag caacttcgac ctggccgagg atgccaaact
     2821 gcagctgagc aaggacacct acgacgacga cctggacaac ctgctggccc agatcggcga
     2881 ccagtacgcc gacctgtttc tggccgccaa gaacctgtcc gacgccatcc tgctgagcga
     2941 catcctgaga gtgaacaccg agatcaccaa ggcccccctg agcgcctcta tgatcaagag
     3001 atacgacgag caccaccagg acctgaccct gctgaaagct ctcgtgcggc agcagctgcc
     3061 tgagaagtac aaagagattt tcttcgacca gagcaagaac ggctacgccg gctacattga
     3121 cggcggagcc agccaggaag agttctacaa gttcatcaag cccatcctgg aaaagatgga
     3181 cggcaccgag gaactgctcg tgaagctgaa cagagaggac ctgctgcgga agcagcggac
     3241 cttcgacaac ggcagcatcc cccaccagat ccacctggga gagctgcacg ccattctgcg
     3301 gcggcaggaa gatttttacc cattcctgaa ggacaaccgg gaaaagatcg agaagatcct
     3361 gaccttccgc atcccctact acgtgggccc tctggccagg ggaaacagca gattcgcctg
     3421 gatgaccaga aagagcgagg aaaccatcac cccctggaac ttcgaggaag tggtggacaa
     3481 gggcgcttcc gcccagagct tcatcgagcg gatgaccaac ttcgataaga acctgcccaa
     3541 cgagaaggtg ctgcccaagc acagcctgct gtacgagtac ttcaccgtgt ataacgagct
     3601 gaccaaagtg aaatacgtga ccgagggaat gagaaagccc gccttcctga gcggcgagca
     3661 gaaaaaggcc atcgtggacc tgctgttcaa gaccaaccgg aaagtgaccg tgaagcagct
     3721 gaaagaggac tacttcaaga aaatcgagtg cttcgactcc gtggaaatct ccggcgtgga
     3781 agatcggttc aacgcctccc tgggcacata ccacgatctg ctgaaaatta tcaaggacaa
     3841 ggacttcctg gacaatgagg aaaacgagga cattctggaa gatatcgtgc tgaccctgac
     3901 actgtttgag gacagagaga tgatcgagga acggctgaaa acctatgccc acctgttcga
     3961 cgacaaagtg atgaagcagc tgaagcggcg gagatacacc ggctggggca ggctgagccg
     4021 gaagctgatc aacggcatcc gggacaagca gtccggcaag acaatcctgg atttcctgaa
     4081 gtccgacggc ttcgccaaca gaaacttcat gcagctgatc cacgacgaca gcctgacctt
     4141 taaagaggac atccagaaag cccaggtgtc cggccagggc gatagcctgc acgagcacat
     4201 tgccaatctg gccggcagcc ccgccattaa gaagggcatc ctgcagacag tgaaggtggt
     4261 ggacgagctc gtgaaagtga tgggccggca caagcccgag aacatcgtga tcgaaatggc
     4321 cagagagaac cagaccaccc agaagggaca gaagaacagc cgcgagagaa tgaagcggat
     4381 cgaagagggc atcaaagagc tgggcagcca gatcctgaaa gaacaccccg tggaaaacac
     4441 ccagctgcag aacgagaagc tgtacctgta ctacctgcag aatgggcggg atatgtacgt
     4501 ggaccaggaa ctggacatca accggctgtc cgactacgat gtggaccata tcgtgcctca
     4561 gagctttctg aaggacgact ccatcgacaa caaggtgctg accagaagcg acaagaaccg
     4621 gggcaagagc gacaacgtgc cctccgaaga ggtcgtgaag aagatgaaga actactggcg
     4681 gcagctgctg aacgccaagc tgattaccca gagaaagttc gacaatctga ccaaggccga
     4741 gagaggcggc ctgagcgaac tggataaggc cggcttcatc aagagacagc tggtggaaac
     4801 ccggcagatc acaaagcacg tggcacagat cctggactcc cggatgaaca ctaagtacga
     4861 cgagaatgac aagctgatcc gggaagtgaa agtgatcacc ctgaagtcca agctggtgtc
     4921 cgatttccgg aaggatttcc agttttacaa agtgcgcgag atcaacaact accaccacgc
     4981 ccacgacgcc tacctgaacg ccgtcgtggg aaccgccctg atcaaaaagt accctaagct
     5041 ggaaagcgag ttcgtgtacg gcgactacaa ggtgtacgac gtgcggaaga tgatcgccaa
     5101 gagcgagcag gaaatcggca aggctaccgc caagtacttc ttctacagca acatcatgaa
     5161 ctttttcaag accgagatta ccctggccaa cggcgagatc cggaagcggc ctctgatcga
     5221 gacaaacggc gaaaccgggg agatcgtgtg ggataagggc cgggattttg ccaccgtgcg
     5281 gaaagtgctg agcatgcccc aagtgaatat cgtgaaaaag accgaggtgc agacaggcgg
     5341 cttcagcaaa gagtctatcc tgcccaagag gaacagcgat aagctgatcg ccagaaagaa
     5401 ggactgggac cctaagaagt acggcggctt cgacagcccc accgtggcct attctgtgct
     5461 ggtggtggcc aaagtggaaa agggcaagtc caagaaactg aagagtgtga aagagctgct
     5521 ggggatcacc atcatggaaa gaagcagctt cgagaagaat cccatcgact ttctggaagc
     5581 caagggctac aaagaagtga aaaaggacct gatcatcaag ctgcctaagt actccctgtt
     5641 cgagctggaa aacggccgga agagaatgct ggcctctgcc ggcgaactgc agaagggaaa
     5701 cgaactggcc ctgccctcca aatatgtgaa cttcctgtac ctggccagcc actatgagaa
     5761 gctgaagggc tcccccgagg ataatgagca gaaacagctg tttgtggaac agcacaagca
     5821 ctacctggac gagatcatcg agcagatcag cgagttctcc aagagagtga tcctggccga
     5881 cgctaatctg gacaaagtgc tgtccgccta caacaagcac cgggataagc ccatcagaga
     5941 gcaggccgag aatatcatcc acctgtttac cctgaccaat ctgggagccc ctgccgcctt
     6001 caagtacttt gacaccacca tcgaccggaa gaggtacacc agcaccaaag aggtgctgga
     6061 cgccaccctg atccaccaga gcatcaccgg cctgtacgag acacggatcg acctgtctca
     6121 gctgggaggc gacagcccca agaagaagag aaaggtggag gccagctaag attcgatctt
     6181 tttccctctg ccaaaaatta tggggacatc atgaagcccc ttgagcatct gacttctggc
     6241 taataaagga aatttatttt cattgcaata gtgtgttgga attttttgtg tctctcactc
     6301 ggtagagtcg acgtcaaggt cgggcaggaa gagggcctat ttcccatgat tccttcatat
     6361 ttgcatatac gatacaaggc tgttagagag ataattagaa ttaatttgac tgtaaacaca
     6421 aagatattag tacaaaatac gtgacgtaga aagtaataat ttcttgggta gtttgcagtt
     6481 ttaaaattat gttttaaaat ggactatcat atgcttaccg taacttgaaa gtatttcgat
     6541 ttcttggctt tatatatctt gtggaaagga cgaaacaccg agcatagcaa gttaaaataa
     6601 ggctagtccg ttatcaactt gaaaaagtgg caccgagtcg gtgctttttt ttctagaggc
     6661 cgcaggaact cgacctcgag acaaatggca gtattcatcc acaattttaa aagaaaaggg
     6721 gggattgggg ggtacagtgc aggggaaaga atagtagaca taatagcaac agacatacaa
     6781 actaaagaat tacaaaaaca aattacaaaa attcaaaatt ttcgggttta ttacagggac
     6841 agcagagatc cactttggcc gcggctcgag ggggttgggg ttgcgccttt tccaaggcag
     6901 ccctgggttt gcgcagggac gcggctgctc tgggcgtggt tccgggaaac gcagcggcgc
     6961 cgaccctggg tctcgcacat tcttcacgtc cgttcgcagc gtcacccgga tcttcgccgc
     7021 tacccttgtg ggccccccgg cgacgcttcc tgctccgccc ctaagtcggg aaggttcctt
     7081 gcggttcgcg gcgtgccgga cgtgacaaac ggaagccgca cgtctcacta gtaccctcgc
     7141 agacggacag cgccagggag caatggcagc gcgccgaccg cgatgggctg tggccaatag
     7201 cggctgctca gcagggcgcg ccgagagcag cggccgggaa ggggcggtgc gggaggcggg
     7261 gtgtggggcg gtagtgtggg ccctgttcct gcccgcgcgg tgttccgcat tctgcaagcc
     7321 tccggagcgc acgtcggcag tcggctccct cgttgaccga atcaccgacc tctctcccca
     7381 gggggatcca ccggagctta ccatgaccga gtacaagccc acggtgcgcc tcgccacccg
     7441 cgacgacgtc cccagggccg tacgcaccct cgccgccgcg ttcgccgact accccgccac
     7501 gcgccacacc gtcgatccgg accgccacat cgagcgggtc accgagctgc aagaactctt
     7561 cctcacgcgc gtcgggctcg acatcggcaa ggtgtgggtc gcggacgacg gcgccgcggt
     7621 ggcggtctgg accacgccgg agagcgtcga agcgggggcg gtgttcgccg agatcggccc
     7681 gcgcatggcc gagttgagcg gttcccggct ggccgcgcag caacagatgg aaggcctcct
     7741 ggcgccgcac cggcccaagg agcccgcgtg gttcctggcc accgtcggcg tctcgcccga
     7801 ccaccagggc aagggtctgg gcagcgccgt cgtgctcccc ggagtggagg cggccgagcg
     7861 cgccggggtg cccgccttcc tggagacctc cgcgccccgc aacctcccct tctacgagcg
     7921 gctcggcttc accgtcaccg ccgacgtcga ggtgcccgaa ggaccgcgca cctggtgcat
     7981 gacccgcaag cccggtgcct gacgcccgcc ccacgacccg cagcgcccga ccgaaaggag
     8041 cgcacgaccc catgcatcgg tacctttaag accaatgact tacaaggcag ctgtagatct
     8101 tagccacttt ttaaaagaaa aggggggact ggaagggcta attcactccc aacgaagaca
     8161 agatctgctt tttgcttgta ctgggtctct ctggttagac cagatctgag cctgggagct
     8221 ctctggctaa ctagggaacc cactgcttaa gcctcaataa agcttgcctt gagtgcttca
     8281 agtagtgtgt gcccgtctgt tgtgtgactc tggtaactag agatccctca gaccctttta
     8341 gtcagtgtgg aaaatctcta gcagtagtag ttcatgtcat cttattattc agtatttata
     8401 acttgcaaag aaatgaatat cagagagtga gaggaacttg tttattgcag cttataatgg
     8461 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
     8521 tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctggctct agctatcccg
     8581 cccctaactc cgcccatccc gcccctaact ccgcccagtt ccgcccattc tccgccccat
     8641 ggctgactaa ttttttttat ttatgcagag gccgaggccg cctcggcctc tgagctattc
     8701 cagaagtagt gaggaggctt ttttggaggc ctagggacgt acccaattcg ccctatagtg
     8761 agtcgtatta cgcgcgctca ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg
     8821 gcgttaccca acttaatcgc cttgcagcac atcccccttt cgccagctgg cgtaatagcg
     8881 aagaggcccg caccgatcgc ccttcccaac agttgcgcag cctgaatggc gaatgggacg
     8941 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     9001 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     9061 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     9121 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     9181 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     9241 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     9301 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     9361 cgaattttaa caaaatatta acgcttacaa tttaggtggc acttttcggg gaaatgtgcg
     9421 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     9481 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     9541 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     9601 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     9661 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     9721 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     9781 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     9841 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     9901 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     9961 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
    10021 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
    10081 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
    10141 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
    10201 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
    10261 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
    10321 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
    10381 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
    10441 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
    10501 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
    10561 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
    10621 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
    10681 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
    10741 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
    10801 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
    10861 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
    10921 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
    10981 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
    11041 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
    11101 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
    11161 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
    11221 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
    11281 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa
    11341 accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga
    11401 ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc attaggcacc
    11461 ccaggcttta cactttggta atgactccaa cttattgata gtgttttatg ttcagataat
    11521 gcccgatgac tttgtcatgc agctccaccg attttgagaa cgacagcgac ttccgtccca
    11581 gccgtgccag gtgctgcctc agattcaggt tatgccgctc aattcgctgc gtatatcgct
    11641 tgctgattac gtgcagcttt cccttcaggc gggattcata cagcggccag ccatccgtca
    11701 tccatatcac cacgtcaaag ggtgacagca ggctcataag acgccccagc gtcgccatag
    11761 tgcgttcacc gaatacgtgc gcaacaaccg tcttccggag actgtcatac gcgtaaaaca
    11821 gccagcgctg gcgcgattta gccccgacat agccccactg ttcgtccatt tccgcgcaga
    11881 cgatgacgtc actgcccggc tgtatgcgcg aggttaccga ctgcggcctg agttttttaa
    11941 gtgacgtaaa atcgtgttga ggccaacgcc cataatgcgg gctgttgccc ggcatccaac
    12001 gccattcatg gccatatcaa tgattttctg gtgcgtaccg ggttgagaag cggtgtaagt
    12061 gaactgcagt tgccatgttt tacggcagtg agagcagaga tagcgctgat gtccggcggt
    12121 gcttttgccg ttacgcacca ccccgtcagt agctgaacag gagggacagc tgatagaaac
    12181 agaagccact ggagcacctc aaaaacacca tcatacacta aatcagtaag ttggcagcat
    12241 caccttacac tttatgcttc cggctcgtat gttgtgtgga attgtgagcg gataacaatt
    12301 tcacacagga aacagctatg accatgatta cgccaagcgc gcaattaacc ctcactaaag
    12361 ggaacaaaag ctggagctgc aagctt
//
