LOCUS       sequence-298676-       11316 bp    DNA     circular SYN 14-APR-2025
DEFINITION  A guide RNA targeting TMPRSS2 in a lentiviral plasmid co-expressing 
            Cas9 and TagBFP2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11316)
  TITLE     CRISPR plasmids to knock out SARS-CoV-2 host genes
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 11316)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: TMPRSS2 g1 + Cas9
FEATURES             Location/Qualifiers
     source          1..11316
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     132..151
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     385..402
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    519..540
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(555..560,561..578,579..585)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    593..609
                     /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     598..620
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     617..633
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     617..633
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     652..672
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        654..672
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        700..926
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             927..1107
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1154..1279
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1772..2005
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     promoter        complement(2183..2431)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(2241..2260)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(2411..2431)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     misc_feature    2438..2555
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2624..2835
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      2855..2864
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2864..2887
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2894..2914
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2939..7039
                     /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             2939..7039
                     /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             7040..7087
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7103..7159
                     /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             7160..7861
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK
                     LVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC
                     DLPSKLGHKLN"
     misc_feature    7868..8456
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7921..7941)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8339..8350)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             8528..8761
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    8839..8960
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8876..8895)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8930..8949
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      9000..9135
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(9155..9174)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(9156..9174)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(9184..9201)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(9184..9200)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9193..9215)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      9342..9797
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9429..9448)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9639..9660
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        9823..9927
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(9928..9996,9997..10788)
                     /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(10146..10165)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(10959..11316,1..231)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
       61 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
      121 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
      181 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
      241 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
      301 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct
      361 cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgccca
      421 atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg
      481 tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta gctcactcat
      541 taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc
      601 ggataacaat ttcacacagg aaacagctat gaccatgatt acgccaagcg cgcaattaac
      661 cctcactaaa gggaacaaaa gctggagctg caagcttaat gtagtcttat gcaatactct
      721 tgtagtcttg caacatggta acgatgagtt agcaacatgc cttacaagga gagaaaaagc
      781 accgtgcatg ccgattggtg gaagtaaggt ggtacgatcg tgccttatta ggaaggcaac
      841 agacgggtct gacatggatt ggacgaacca ctgaattgcc gcattgcaga gatattgtat
      901 ttaagtgcct agctcgatac ataaacgggt ctctctggtt agaccagatc tgagcctggg
      961 agctctctgg ctaactaggg aacccactgc ttaagcctca ataaagcttg ccttgagtgc
     1021 ttcaagtagt gtgtgcccgt ctgttgtgtg actctggtaa ctagagatcc ctcagaccct
     1081 tttagtcagt gtggaaaatc tctagcagtg gcgcccgaac agggacttga aagcgaaagg
     1141 gaaaccagag gagctctctc gacgcaggac tcggcttgct gaagcgcgca cggcaagagg
     1201 cgaggggcgg cgactggtga gtacgccaaa aattttgact agcggaggct agaaggagag
     1261 agatgggtgc gagagcgtca gtattaagcg ggggagaatt agatcgcgat gggaaaaaat
     1321 tcggttaagg ccagggggaa agaaaaaata taaattaaaa catatagtat gggcaagcag
     1381 ggagctagaa cgattcgcag ttaatcctgg cctgttagaa acatcagaag gctgtagaca
     1441 aatactggga cagctacaac catcccttca gacaggatca gaagaactta gatcattata
     1501 taatacagta gcaaccctct attgtgtgca tcaaaggata gagataaaag acaccaagga
     1561 agctttagac aagatagagg aagagcaaaa caaaagtaag accaccgcac agcaagcggc
     1621 cgctgatctt cagacctgga ggaggagata tgagggacaa ttggagaagt gaattatata
     1681 aatataaagt agtaaaaatt gaaccattag gagtagcacc caccaaggca aagagaagag
     1741 tggtgcagag agaaaaaaga gcagtgggaa taggagcttt gttccttggg ttcttgggag
     1801 cagcaggaag cactatgggc gcagcgtcaa tgacgctgac ggtacaggcc agacaattat
     1861 tgtctggtat agtgcagcag cagaacaatt tgctgagggc tattgaggcg caacagcatc
     1921 tgttgcaact cacagtctgg ggcatcaagc agctccaggc aagaatcctg gctgtggaaa
     1981 gatacctaaa ggatcaacag ctcctgggga tttggggttg ctctggaaaa ctcgccattt
     2041 gtctcaagat ctagaattca aaaaaagcac cgactcggtg ccactttgct gtttccagca
     2101 aagttgataa cggactagcc ttatttaaac ttgctatgct gtttccagca tagctcttaa
     2161 accgcaaatg ccgtccaatg ccggtgtttc gtcctttcca caagatatat aaagccaaga
     2221 aatcgaaata ctttcaagtt acggtaagca tatgatagtc cattttaaaa cataatttta
     2281 aaactgcaaa ctacccaaga aattattact ttctacgtca cgtattttgt actaatatct
     2341 ttgtgtttac agtcaaatta attctaatta tctctctaac agccttgtat cgtatatgca
     2401 aatatgaagg aatcatggga aataggccct cccacaattt taaaagaaaa ggggggattg
     2461 gggggtacag tgcaggggaa agaatagtag acataatagc aacagacata caaactaaag
     2521 aattacaaaa acaaattaca aaaattcaaa attttcgggt ttattacagg gacagcagag
     2581 atccactttg gcgccggctc gagtggctcc ggtgcccgtc agtgggcaga gcgcacatcg
     2641 cccacagtcc ccgagaagtt ggggggaggg gtcggcaatt gaaccggtgc ctagagaagg
     2701 tggcgcgggg taaactggga aagtgatgtc gtgtactggc tccgcctttt tcccgagggt
     2761 gggggagaac cgtatataag tgcagtagtc gccgtgaacg ttctttttcg caacgggttt
     2821 gccgccagaa cacaggtgtc gtgacgcggg atccgccacc atggattaca aagacgatga
     2881 cgataagatg gccccaaaga agaagcggaa ggtcggtatc cacggagtcc cagcagccga
     2941 caagaagtac agcatcggcc tggacatcgg caccaactct gtgggctggg ccgtgatcac
     3001 cgacgagtac aaggtgccca gcaagaaatt caaggtgctg ggcaacaccg accggcacag
     3061 catcaagaag aacctgatcg gagccctgct gttcgacagc ggcgaaacag ccgaggccac
     3121 ccggctgaag agaaccgcca gaagaagata caccagacgg aagaaccgga tctgctatct
     3181 gcaagagatc ttcagcaacg agatggccaa ggtggacgac agcttcttcc acagactgga
     3241 agagtccttc ctggtggaag aggataagaa gcacgagcgg caccccatct tcggcaacat
     3301 cgtggacgag gtggcctacc acgagaagta ccccaccatc taccacctga gaaagaaact
     3361 ggtggacagc accgacaagg ccgacctgcg gctgatctat ctggccctgg cccacatgat
     3421 caagttccgg ggccacttcc tgatcgaggg cgacctgaac cccgacaaca gcgacgtgga
     3481 caagctgttc atccagctgg tgcagaccta caaccagctg ttcgaggaaa accccatcaa
     3541 cgccagcggc gtggacgcca aggccatcct gtctgccaga ctgagcaaga gcagacggct
     3601 ggaaaatctg atcgcccagc tgcccggcga gaagaagaat ggcctgttcg gaaacctgat
     3661 tgccctgagc ctgggcctga cccccaactt caagagcaac ttcgacctgg ccgaggatgc
     3721 caaactgcag ctgagcaagg acacctacga cgacgacctg gacaacctgc tggcccagat
     3781 cggcgaccag tacgccgacc tgtttctggc cgccaagaac ctgtccgacg ccatcctgct
     3841 gagcgacatc ctgagagtga acaccgagat caccaaggcc cccctgagcg cctctatgat
     3901 caagagatac gacgagcacc accaggacct gaccctgctg aaagctctcg tgcggcagca
     3961 gctgcctgag aagtacaaag agattttctt cgaccagagc aagaacggct acgccggcta
     4021 cattgacggc ggagccagcc aggaagagtt ctacaagttc atcaagccca tcctggaaaa
     4081 gatggacggc accgaggaac tgctcgtgaa gctgaacaga gaggacctgc tgcggaagca
     4141 gcggaccttc gacaacggca gcatccccca ccagatccac ctgggagagc tgcacgccat
     4201 tctgcggcgg caggaagatt tttacccatt cctgaaggac aaccgggaaa agatcgagaa
     4261 gatcctgacc ttccgcatcc cctactacgt gggccctctg gccaggggaa acagcagatt
     4321 cgcctggatg accagaaaga gcgaggaaac catcaccccc tggaacttcg aggaagtggt
     4381 ggacaagggc gcttccgccc agagcttcat cgagcggatg accaacttcg ataagaacct
     4441 gcccaacgag aaggtgctgc ccaagcacag cctgctgtac gagtacttca ccgtgtataa
     4501 cgagctgacc aaagtgaaat acgtgaccga gggaatgaga aagcccgcct tcctgagcgg
     4561 cgagcagaaa aaggccatcg tggacctgct gttcaagacc aaccggaaag tgaccgtgaa
     4621 gcagctgaaa gaggactact tcaagaaaat cgagtgcttc gactccgtgg aaatctccgg
     4681 cgtggaagat cggttcaacg cctccctggg cacataccac gatctgctga aaattatcaa
     4741 ggacaaggac ttcctggaca atgaggaaaa cgaggacatt ctggaagata tcgtgctgac
     4801 cctgacactg tttgaggaca gagagatgat cgaggaacgg ctgaaaacct atgcccacct
     4861 gttcgacgac aaagtgatga agcagctgaa gcggcggaga tacaccggct ggggcaggct
     4921 gagccggaag ctgatcaacg gcatccggga caagcagtcc ggcaagacaa tcctggattt
     4981 cctgaagtcc gacggcttcg ccaacagaaa cttcatgcag ctgatccacg acgacagcct
     5041 gacctttaaa gaggacatcc agaaagccca ggtgtccggc cagggcgata gcctgcacga
     5101 gcacattgcc aatctggccg gcagccccgc cattaagaag ggcatcctgc agacagtgaa
     5161 ggtggtggac gagctcgtga aagtgatggg ccggcacaag cccgagaaca tcgtgatcga
     5221 aatggccaga gagaaccaga ccacccagaa gggacagaag aacagccgcg agagaatgaa
     5281 gcggatcgaa gagggcatca aagagctggg cagccagatc ctgaaagaac accccgtgga
     5341 aaacacccag ctgcagaacg agaagctgta cctgtactac ctgcagaatg ggcgggatat
     5401 gtacgtggac caggaactgg acatcaaccg gctgtccgac tacgatgtgg accatatcgt
     5461 gcctcagagc tttctgaagg acgactccat cgacaacaag gtgctgacca gaagcgacaa
     5521 gaaccggggc aagagcgaca acgtgccctc cgaagaggtc gtgaagaaga tgaagaacta
     5581 ctggcggcag ctgctgaacg ccaagctgat tacccagaga aagttcgaca atctgaccaa
     5641 ggccgagaga ggcggcctga gcgaactgga taaggccggc ttcatcaaga gacagctggt
     5701 ggaaacccgg cagatcacaa agcacgtggc acagatcctg gactcccgga tgaacactaa
     5761 gtacgacgag aatgacaagc tgatccggga agtgaaagtg atcaccctga agtccaagct
     5821 ggtgtccgat ttccggaagg atttccagtt ttacaaagtg cgcgagatca acaactacca
     5881 ccacgcccac gacgcctacc tgaacgccgt cgtgggaacc gccctgatca aaaagtaccc
     5941 taagctggaa agcgagttcg tgtacggcga ctacaaggtg tacgacgtgc ggaagatgat
     6001 cgccaagagc gagcaggaaa tcggcaaggc taccgccaag tacttcttct acagcaacat
     6061 catgaacttt ttcaagaccg agattaccct ggccaacggc gagatccgga agcggcctct
     6121 gatcgagaca aacggcgaaa ccggggagat cgtgtgggat aagggccggg attttgccac
     6181 cgtgcggaaa gtgctgagca tgccccaagt gaatatcgtg aaaaagaccg aggtgcagac
     6241 aggcggcttc agcaaagagt ctatcctgcc caagaggaac agcgataagc tgatcgccag
     6301 aaagaaggac tgggacccta agaagtacgg cggcttcgac agccccaccg tggcctattc
     6361 tgtgctggtg gtggccaaag tggaaaaggg caagtccaag aaactgaaga gtgtgaaaga
     6421 gctgctgggg atcaccatca tggaaagaag cagcttcgag aagaatccca tcgactttct
     6481 ggaagccaag ggctacaaag aagtgaaaaa ggacctgatc atcaagctgc ctaagtactc
     6541 cctgttcgag ctggaaaacg gccggaagag aatgctggcc tctgccggcg aactgcagaa
     6601 gggaaacgaa ctggccctgc cctccaaata tgtgaacttc ctgtacctgg ccagccacta
     6661 tgagaagctg aagggctccc ccgaggataa tgagcagaaa cagctgtttg tggaacagca
     6721 caagcactac ctggacgaga tcatcgagca gatcagcgag ttctccaaga gagtgatcct
     6781 ggccgacgct aatctggaca aagtgctgtc cgcctacaac aagcaccggg ataagcccat
     6841 cagagagcag gccgagaata tcatccacct gtttaccctg accaatctgg gagcccctgc
     6901 cgccttcaag tactttgaca ccaccatcga ccggaagagg tacaccagca ccaaagaggt
     6961 gctggacgcc accctgatcc accagagcat caccggcctg tacgagacac ggatcgacct
     7021 gtctcagctg ggaggcgaca agcgtcctgc tgctactaag aaagctggtc aagctaagaa
     7081 aaagaaagct agcggcagcg gcgccaccaa cttcagcctg ctgaagcagg ccggcgacgt
     7141 ggaggagaac cccggcccca tgagcgagct gattaaggag aacatgcaca tgaagctgta
     7201 catggagggc accgtggaca accatcactt caagtgcaca tccgagggcg aaggcaagcc
     7261 ctacgagggc acccagacca tgagaatcaa ggtggtcgag ggcggccctc tccccttcgc
     7321 cttcgacatc ctggctacta gcttcctcta cggcagcaag accttcatca accacaccca
     7381 gggcatcccc gacttcttca agcagtcctt ccctgagggc ttcacatggg agagagtcac
     7441 cacatacgaa gacgggggcg tgctgaccgc tacccaggac accagcctcc aggacggctg
     7501 cctcatctac aacgtcaaga tcagaggggt gaacttcaca tccaacggcc ctgtgatgca
     7561 gaagaaaaca ctcggctggg aggccttcac cgaaacgctg taccccgctg acggcggcct
     7621 ggaaggcaga aacgacatgg ccctgaagct cgtgggcggg agccatctga tcgcaaacgc
     7681 caagaccaca tatagatcca agaaacccgc taagaacctc aagatgcctg gcgtctacta
     7741 tgtggactac agactggaaa gaatcaagga ggccaacaac gagacctacg tcgagcagca
     7801 cgaggtggca gtggccagat actgcgacct ccctagcaaa ctggggcaca agcttaattg
     7861 agtcgacaat caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta
     7921 tgttgctcct tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc
     7981 ttcccgtatg gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga
     8041 ggagttgtgg cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac
     8101 ccccactggt tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc
     8161 cctccctatt gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc
     8221 tcggctgttg ggcactgaca attccgtggt gttgtcgggg aaatcatcgt cctttccttg
     8281 gctgctcgcc tgtgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc
     8341 ggccctcaat ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc
     8401 gcgtcttcgc cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcgtcg
     8461 actttaagac caatgactta caaggcagct gtagatctta gccacttttt aaaagaaaag
     8521 gggggactgg aagggctaat tcactcccaa cgaagacaag atctgctttt tgcttgtact
     8581 gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact agggaaccca
     8641 ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg
     8701 tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa aatctctagc
     8761 agtacgtata gtagttcatg tcatcttatt attcagtatt tataacttgc aaagaaatga
     8821 atatcagaga gtgagaggaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     8881 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     8941 aaactcatca atgtatctta tcatgtctgg ctctagctat cccgccccta actccgccca
     9001 tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt
     9061 ttatttatgc agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag
     9121 gcttttttgg aggcctaggg acgtacccaa ttcgccctat agtgagtcgt attacgcgcg
     9181 ctcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa
     9241 tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga
     9301 tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg gacgcgccct gtagcggcgc
     9361 attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct
     9421 agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg
     9481 tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga
     9541 ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt
     9601 ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg
     9661 aacaacactc aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc
     9721 ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat
     9781 attaacgctt acaatttagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     9841 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     9901 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     9961 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
    10021 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
    10081 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
    10141 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
    10201 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
    10261 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
    10321 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
    10381 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
    10441 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
    10501 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
    10561 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
    10621 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
    10681 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
    10741 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
    10801 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
    10861 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
    10921 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
    10981 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
    11041 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
    11101 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
    11161 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
    11221 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
    11281 gggggttcgt gcacacagcc cagcttggag cgaacg
//
