LOCUS       sequence-428808-       13842 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral expression vector for an inducible Cas9-P2A-BFP with two 
            empty sgRNA cloning sites.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13842)
  TITLE     Selectable lentivectors delivering dual gRNAs and destabilized Cas9
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 13842)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pDual_dsCas9_BFP
FEATURES             Location/Qualifiers
     source          1..13842
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(47..66)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        238..617
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..817
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     772..792
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             835..1015
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1062..1187
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1680..1913
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2098..2142
                     /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    2440..2557
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2608..2848
                     /label=U6 promoter
                     /note="Truncated RNA polymerase III promoter for human U6 
                     snRNA used in pSIREN vectors such as 
                     pSIREN-RetroQ-ZsGreen1. Expected transcription start site 
                     is 8bp downstream of this feature."
     misc_RNA        2860..2935
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        3026..3266
                     /label=U6 promoter
                     /note="Truncated RNA polymerase III promoter for human U6 
                     snRNA used in pSIREN vectors such as 
                     pSIREN-RetroQ-ZsGreen1. Expected transcription start site 
                     is 8bp downstream of this feature."
     misc_RNA        3274..3349
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        3411..3622
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      3636..3645
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3642..3644,3645..3965)
                     /codon_start=1
                     /product="destabilization domain that can be stabilized by 
                     Shield1 in the ProteoTuner(TM) system"
                     /label=DD
                     /note="L106P mutant of FKBP12"
                     /translation="MGVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKVDSSRDRNK
                     PFKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVEL
                     LKPE"
     CDS             3975..8078
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             3975..8078
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             8079..8126
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             8127..8150
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             8160..8216
                     /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             8217..8918
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK
                     LVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC
                     DLPSKLGHKLN"
     misc_feature    8934..9522
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8987..9007)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(9405..9416)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             9594..9827
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(9853..9870)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9859..10083
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      10129..10557
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10216..10235)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10426..10447
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10566..10586)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10571..10900
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      10751..10886
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10813..10832
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10948..10995
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             11014..11388
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    11518..11639
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11555..11574)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11609..11628
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11688..11704)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11688..11704)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11701..11723)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11712..11728
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(11736..11742,11743..11760,11761..11766))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11781..11802
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11919..11936)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(12090..12678)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(12170..12189)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12849..13640,13641..13709))
                     /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     13472..13491
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(13710..13814)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
      241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
      301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct
      841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
      901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
      961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
     1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
     1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
     1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
     1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
     1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
     1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
     1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
     1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca
     1501 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg
     1561 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga
     1621 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata
     1681 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg
     1741 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg
     1801 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag
     1861 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt
     1921 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt
     1981 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt
     2041 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag
     2101 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata
     2161 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta
     2221 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta
     2281 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa
     2341 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt
     2401 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat
     2461 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa
     2521 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag
     2581 agatccagtt tggttaatta aggtaccgag ggcctatttc ccatgattcc ttcatatttg
     2641 catatacgat acaaggctgt tagagagata attagaatta atttgactgt aaacacaaag
     2701 atattagtac aaaatacgtg acgtagaaag taataatttc ttgggtagtt tgcagtttta
     2761 aaattatgtt ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta tttcgatttc
     2821 ttggctttat atatcttgtg gaaaggacga aacctgcagg ttttagagct agaaatagca
     2881 agttaaaata aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc ggtgcttttt
     2941 tgttttagag ctagaaatag caagttaaaa taaggctagt ccgtttttag cgcgtgcgcc
     3001 aattctgcag acaaatggct ctagagaggg cctatttccc atgattcctt catatttgca
     3061 tatacgatac aaggctgtta gagagataat tggaattaat ttgactgtaa acacaaagat
     3121 attagtacaa aatacgtgac gtagaaagta ataatttctt gggtagtttg cagttttaaa
     3181 attatgtttt aaaatggact atcatatgct taccgtaact tgaaagtatt tcgatttctt
     3241 ggctttatat atcttgtgga aaggaccgtc tctgttttag agctagaaat agcaagttaa
     3301 aataaggcta gtccgttatc aacttgaaaa agtggcaccg agtcggtgct tttttgaatt
     3361 cgctagctag gtcttgaaag gagtgggaat tggctccggt gcccgtcagt gggcagagcg
     3421 cacatcgccc acagtccccg agaagttggg gggaggggtc ggcaattgat ccggtgccta
     3481 gagaaggtgg cgcggggtaa actgggaaag tgatgtcgtg tactggctcc gcctttttcc
     3541 cgagggtggg ggagaaccgt atataagtgc agtagtcgcc gtgaacgttc tttttcgcaa
     3601 cgggtttgcc gccagaacac aggaccggtt ctagagccac catgggagtg caggtggaaa
     3661 ccatctcccc aggtgacggg cgcaccttcc ccaagcgcgg ccagacctgt gtggtgcact
     3721 acaccgggat gcttgaagat ggaaagaaag tcgattcctc ccgggacaga aacaagccct
     3781 ttaagtttat gctaggcaag caggaggtga tccgaggctg ggaagaaggg gttgcccaga
     3841 tgagtgtggg tcagagagcc aaactgacta tatctccaga ttatgcctat ggtgccactg
     3901 ggcacccagg catcatccca ccacatgcca ctctcgtctt cgatgtggag cttctaaaac
     3961 cggaaccggc tagcatggac aagaagtaca gcatcggcct ggacatcggc accaactctg
     4021 tgggctgggc cgtgatcacc gacgagtaca aggtgcccag caagaaattc aaggtgctgg
     4081 gcaacaccga ccggcacagc atcaagaaga acctgatcgg agccctgctg ttcgacagcg
     4141 gcgaaacagc cgaggccacc cggctgaaga gaaccgccag aagaagatac accagacgga
     4201 agaaccggat ctgctatctg caagagatct tcagcaacga gatggccaag gtggacgaca
     4261 gcttcttcca cagactggaa gagtccttcc tggtggaaga ggataagaag cacgagcggc
     4321 accccatctt cggcaacatc gtggacgagg tggcctacca cgagaagtac cccaccatct
     4381 accacctgag aaagaaactg gtggacagca ccgacaaggc cgacctgcgg ctgatctatc
     4441 tggccctggc ccacatgatc aagttccggg gccacttcct gatcgagggc gacctgaacc
     4501 ccgacaacag cgacgtggac aagctgttca tccagctggt gcagacctac aaccagctgt
     4561 tcgaggaaaa ccccatcaac gccagcggcg tggacgccaa ggccatcctg tctgccagac
     4621 tgagcaagag cagacggctg gaaaatctga tcgcccagct gcccggcgag aagaagaatg
     4681 gcctgttcgg aaacctgatt gccctgagcc tgggcctgac ccccaacttc aagagcaact
     4741 tcgacctggc cgaggatgcc aaactgcagc tgagcaagga cacctacgac gacgacctgg
     4801 acaacctgct ggcccagatc ggcgaccagt acgccgacct gtttctggcc gccaagaacc
     4861 tgtccgacgc catcctgctg agcgacatcc tgagagtgaa caccgagatc accaaggccc
     4921 ccctgagcgc ctctatgatc aagagatacg acgagcacca ccaggacctg accctgctga
     4981 aagctctcgt gcggcagcag ctgcctgaga agtacaaaga gattttcttc gaccagagca
     5041 agaacggcta cgccggctac attgacggcg gagccagcca ggaagagttc tacaagttca
     5101 tcaagcccat cctggaaaag atggacggca ccgaggaact gctcgtgaag ctgaacagag
     5161 aggacctgct gcggaagcag cggaccttcg acaacggcag catcccccac cagatccacc
     5221 tgggagagct gcacgccatt ctgcggcggc aggaagattt ttacccattc ctgaaggaca
     5281 accgggaaaa gatcgagaag atcctgacct tccgcatccc ctactacgtg ggccctctgg
     5341 ccaggggaaa cagcagattc gcctggatga ccagaaagag cgaggaaacc atcaccccct
     5401 ggaacttcga ggaagtggtg gacaagggcg cttccgccca gagcttcatc gagcggatga
     5461 ccaacttcga taagaacctg cccaacgaga aggtgctgcc caagcacagc ctgctgtacg
     5521 agtacttcac cgtgtataac gagctgacca aagtgaaata cgtgaccgag ggaatgagaa
     5581 agcccgcctt cctgagcggc gagcagaaaa aggccatcgt ggacctgctg ttcaagacca
     5641 accggaaagt gaccgtgaag cagctgaaag aggactactt caagaaaatc gagtgcttcg
     5701 actccgtgga aatctccggc gtggaagatc ggttcaacgc ctccctgggc acataccacg
     5761 atctgctgaa aattatcaag gacaaggact tcctggacaa tgaggaaaac gaggacattc
     5821 tggaagatat cgtgctgacc ctgacactgt ttgaggacag agagatgatc gaggaacggc
     5881 tgaaaaccta tgcccacctg ttcgacgaca aagtgatgaa gcagctgaag cggcggagat
     5941 acaccggctg gggcaggctg agccggaagc tgatcaacgg catccgggac aagcagtccg
     6001 gcaagacaat cctggatttc ctgaagtccg acggcttcgc caacagaaac ttcatgcagc
     6061 tgatccacga cgacagcctg acctttaaag aggacatcca gaaagcccag gtgtccggcc
     6121 agggcgatag cctgcacgag cacattgcca atctggccgg cagccccgcc attaagaagg
     6181 gcatcctgca gacagtgaag gtggtggacg agctcgtgaa agtgatgggc cggcacaagc
     6241 ccgagaacat cgtgatcgaa atggccagag agaaccagac cacccagaag ggacagaaga
     6301 acagccgcga gagaatgaag cggatcgaag agggcatcaa agagctgggc agccagatcc
     6361 tgaaagaaca ccccgtggaa aacacccagc tgcagaacga gaagctgtac ctgtactacc
     6421 tgcagaatgg gcgggatatg tacgtggacc aggaactgga catcaaccgg ctgtccgact
     6481 acgatgtgga ccatatcgtg cctcagagct ttctgaagga cgactccatc gacaacaagg
     6541 tgctgaccag aagcgacaag aaccggggca agagcgacaa cgtgccctcc gaagaggtcg
     6601 tgaagaagat gaagaactac tggcggcagc tgctgaacgc caagctgatt acccagagaa
     6661 agttcgacaa tctgaccaag gccgagagag gcggcctgag cgaactggat aaggccggct
     6721 tcatcaagag acagctggtg gaaacccggc agatcacaaa gcacgtggca cagatcctgg
     6781 actcccggat gaacactaag tacgacgaga atgacaagct gatccgggaa gtgaaagtga
     6841 tcaccctgaa gtccaagctg gtgtccgatt tccggaagga tttccagttt tacaaagtgc
     6901 gcgagatcaa caactaccac cacgcccacg acgcctacct gaacgccgtc gtgggaaccg
     6961 ccctgatcaa aaagtaccct aagctggaaa gcgagttcgt gtacggcgac tacaaggtgt
     7021 acgacgtgcg gaagatgatc gccaagagcg agcaggaaat cggcaaggct accgccaagt
     7081 acttcttcta cagcaacatc atgaactttt tcaagaccga gattaccctg gccaacggcg
     7141 agatccggaa gcggcctctg atcgagacaa acggcgaaac cggggagatc gtgtgggata
     7201 agggccggga ttttgccacc gtgcggaaag tgctgagcat gccccaagtg aatatcgtga
     7261 aaaagaccga ggtgcagaca ggcggcttca gcaaagagtc tatcctgccc aagaggaaca
     7321 gcgataagct gatcgccaga aagaaggact gggaccctaa gaagtacggc ggcttcgaca
     7381 gccccaccgt ggcctattct gtgctggtgg tggccaaagt ggaaaagggc aagtccaaga
     7441 aactgaagag tgtgaaagag ctgctgggga tcaccatcat ggaaagaagc agcttcgaga
     7501 agaatcccat cgactttctg gaagccaagg gctacaaaga agtgaaaaag gacctgatca
     7561 tcaagctgcc taagtactcc ctgttcgagc tggaaaacgg ccggaagaga atgctggcct
     7621 ctgccggcga actgcagaag ggaaacgaac tggccctgcc ctccaaatat gtgaacttcc
     7681 tgtacctggc cagccactat gagaagctga agggctcccc cgaggataat gagcagaaac
     7741 agctgtttgt ggaacagcac aagcactacc tggacgagat catcgagcag atcagcgagt
     7801 tctccaagag agtgatcctg gccgacgcta atctggacaa agtgctgtcc gcctacaaca
     7861 agcaccggga taagcccatc agagagcagg ccgagaatat catccacctg tttaccctga
     7921 ccaatctggg agcccctgcc gccttcaagt actttgacac caccatcgac cggaagaggt
     7981 acaccagcac caaagaggtg ctggacgcca ccctgatcca ccagagcatc accggcctgt
     8041 acgagacacg gatcgacctg tctcagctgg gaggcgacaa gcgacctgcc gccacaaaga
     8101 aggctggaca ggctaagaag aagaaagatt acaaagacga tgacgataag ggatccggtg
     8161 ccacgaattt ctcactactc aagcaggccg gtgatgtcga ggaaaaccct ggtcctatga
     8221 gcgagctgat taaggagaac atgcacatga agctgtacat ggagggcacc gtggacaacc
     8281 atcacttcaa gtgcacatcc gagggcgaag gcaagcccta cgagggcacc cagaccatga
     8341 gaatcaaggt ggtcgagggc ggccctctcc ccttcgcctt cgacatcctg gctactagct
     8401 tcctctacgg cagcaagacc ttcatcaacc acacccaggg catccccgac ttcttcaagc
     8461 agtccttccc tgagggcttc acatgggaga gagtcaccac atacgaagac gggggcgtgc
     8521 tgaccgctac ccaggacacc agcctccagg acggctgcct catctacaac gtcaagatca
     8581 gaggggtgaa cttcacatcc aacggccctg tgatgcagaa gaaaacactc ggctgggagg
     8641 ccttcaccga aactctgtac cccgctgacg gcggcctgga aggcagaaac gacatggccc
     8701 tgaagctcgt gggcgggagc catctgatcg caaacgccaa gaccacatat agatccaaga
     8761 aacccgctaa gaacctcaag atgcctggcg tctactatgt ggactacaga ctggaaagaa
     8821 tcaaggaggc caacaacgag acctacgtcg agcagcacga ggtggcagtg gccagatact
     8881 gcgacctccc tagcaaactg gggcacaagc ttaattgagg atcctaagtc gacaatcaac
     8941 ctctggatta caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta
     9001 cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt
     9061 tcattttctc ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg
     9121 ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg
     9181 gcattgccac cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca
     9241 cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca
     9301 ctgacaattc cgtggtgttg tcggggaaat catcgtcctt tccttggctg ctcgcctgtg
     9361 ttgccacctg gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag
     9421 cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc
     9481 gccctcagac gagtcggatc tccctttggg ccgcctcccc gcgtcgactt taagaccaat
     9541 gacttacaag gcagctgtag atcttagcca ctttttaaaa gaaaaggggg gactggaagg
     9601 gctaattcac tcccaacgaa gacaagatct gctttttgct tgtactgggt ctctctggtt
     9661 agaccagatc tgagcctggg agctctctgg ctaactaggg aacccactgc ttaagcctca
     9721 ataaagcttg ccttgagtgc ttcaagtagt gtgtgcccgt ctgttgtgtg actctggtaa
     9781 ctagagatcc ctcagaccct tttagtcagt gtggaaaatc tctagcaggg cccgtttaaa
     9841 cccgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc
     9901 ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg
     9961 aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg
    10021 acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggatgcg gtgggctcta
    10081 tggcttctga ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta
    10141 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
    10201 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
    10261 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
    10321 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
    10381 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
    10441 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
    10501 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat
    10561 tctgtggaat gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag
    10621 tatgcaaagc atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc
    10681 agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct
    10741 aactccgccc atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg
    10801 actaattttt tttatttatg cagaggccga ggccgcctct gcctctgagc tattccagaa
    10861 gtagtgagga ggcttttttg gaggcctagg cttttgcaaa aagctcccgg gagcttgtat
    10921 atccattttc ggatctgatc agcacgtgtt gacaattaat catcggcata gtatatcggc
    10981 atagtataat acgacaaggt gaggaactaa accatggcca agttgaccag tgccgttccg
    11041 gtgctcaccg cgcgcgacgt cgccggagcg gtcgagttct ggaccgaccg gctcgggttc
    11101 tcccgggact tcgtggagga cgacttcgcc ggtgtggtcc gggacgacgt gaccctgttc
    11161 atcagcgcgg tccaggacca ggtggtgccg gacaacaccc tggcctgggt gtgggtgcgc
    11221 ggcctggacg agctgtacgc cgagtggtcg gaggtcgtgt ccacgaactt ccgggacgcc
    11281 tccgggccgg ccatgaccga gatcggcgag cagccgtggg ggcgggagtt cgccctgcgc
    11341 gacccggccg gcaactgcgt gcacttcgtg gccgaggagc aggactgaca cgtgctacga
    11401 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac
    11461 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac
    11521 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
    11581 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
    11641 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt
    11701 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
    11761 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
    11821 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
    11881 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
    11941 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
    12001 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
    12061 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
    12121 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
    12181 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
    12241 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
    12301 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
    12361 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
    12421 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
    12481 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
    12541 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
    12601 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
    12661 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
    12721 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
    12781 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
    12841 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
    12901 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
    12961 tctggcccca gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca
    13021 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
    13081 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
    13141 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
    13201 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
    13261 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
    13321 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
    13381 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
    13441 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
    13501 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
    13561 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
    13621 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
    13681 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
    13741 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
    13801 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg ac
//
