LOCUS       sequence-382143-       13401 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expression of spCas9 and non-targeting control sgRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13401)
  AUTHORS   Castagnino P, Kim HW, Lam LKM, Basu D, White EA
  TITLE     Systematic Analysis of IL-1 Cytokine Signaling Suppression by HPV16 
            Oncoproteins.
  JOURNAL   J Virol. 2022 Nov 7:e0132622. doi: 10.1128/jvi.01326-22.
  PUBMED    36342298
REFERENCE   2  (bases 1 to 13401)
  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: "J Virol. 
            2022 Nov 7:e0132622. doi: 10.1128/jvi.01326-22."
COMMENT     Sequence Label: p8267 LentiCRISPR v2 hygro sgNT-2
FEATURES             Location/Qualifiers
     source          1..13401
                     /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        242..621
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        623..821
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     776..796
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             839..1019
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1066..1191
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1684..1917
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2102..2146
                     /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    2444..2561
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2612..2860
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2612..2632
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2783..2802
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2882..2957
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        3019..3230
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      3249..3258
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3255..7358
                     /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             3255..7358
                     /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             7359..7406
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7407..7430
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             7440..7496
                     /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"
     misc_feature    8493..9081
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8546..8566)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8964..8975)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             9153..9386
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(9412..9429)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9418..9642
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      9688..10116
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9775..9794)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9985..10006
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10125..10145)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10130..10459
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      10310..10445
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10372..10391
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10507..10554
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10573..10947
                     /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    11077..11198
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11114..11133)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11168..11187
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11247..11263)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11247..11263)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11260..11282)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11271..11287
                     /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(11295..11301,11302..11319,11320..11325))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11340..11361
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11478..11495)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11649..12237)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11729..11748)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12408..13199,13200..13268))
                     /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     13031..13050
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(13269..13373)
                     /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 acgcgcgcgt
      241 tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt agttcatagc
      301 ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc
      361 aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac gccaataggg
      421 actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt ggcagtacat
      481 caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa atggcccgcc
      541 tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta catctacgta
      601 ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg gcgtggatag
      661 cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg gagtttgttt
      721 tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc attgacgcaa
      781 atgggcggta ggcgtgtacg gtgggaggtc tatataagca gcgcgttttg cctgtactgg
      841 gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact
      901 gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg
      961 tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag
     1021 tggcgcccga acagggactt gaaagcgaaa gggaaaccag aggagctctc tcgacgcagg
     1081 actcggcttg ctgaagcgcg cacggcaaga ggcgaggggc ggcgactggt gagtacgcca
     1141 aaaattttga ctagcggagg ctagaaggag agagatgggt gcgagagcgt cagtattaag
     1201 cgggggagaa ttagatcgcg atgggaaaaa attcggttaa ggccaggggg aaagaaaaaa
     1261 tataaattaa aacatatagt atgggcaagc agggagctag aacgattcgc agttaatcct
     1321 ggcctgttag aaacatcaga aggctgtaga caaatactgg gacagctaca accatccctt
     1381 cagacaggat cagaagaact tagatcatta tataatacag tagcaaccct ctattgtgtg
     1441 catcaaagga tagagataaa agacaccaag gaagctttag acaagataga ggaagagcaa
     1501 aacaaaagta agaccaccgc acagcaagcg gccgctgatc ttcagacctg gaggaggaga
     1561 tatgagggac aattggagaa gtgaattata taaatataaa gtagtaaaaa ttgaaccatt
     1621 aggagtagca cccaccaagg caaagagaag agtggtgcag agagaaaaaa gagcagtggg
     1681 aataggagct ttgttccttg ggttcttggg agcagcagga agcactatgg gcgcagcgtc
     1741 aatgacgctg acggtacagg ccagacaatt attgtctggt atagtgcagc agcagaacaa
     1801 tttgctgagg gctattgagg cgcaacagca tctgttgcaa ctcacagtct ggggcatcaa
     1861 gcagctccag gcaagaatcc tggctgtgga aagataccta aaggatcaac agctcctggg
     1921 gatttggggt tgctctggaa aactcatttg caccactgct gtgccttgga atgctagttg
     1981 gagtaataaa tctctggaac agatttggaa tcacacgacc tggatggagt gggacagaga
     2041 aattaacaat tacacaagct taatacactc cttaattgaa gaatcgcaaa accagcaaga
     2101 aaagaatgaa caagaattat tggaattaga taaatgggca agtttgtgga attggtttaa
     2161 cataacaaat tggctgtggt atataaaatt attcataatg atagtaggag gcttggtagg
     2221 tttaagaata gtttttgctg tactttctat agtgaataga gttaggcagg gatattcacc
     2281 attatcgttt cagacccacc tcccaacccc gaggggaccc gacaggcccg aaggaataga
     2341 agaagaaggt ggagagagag acagagacag atccattcga ttagtgaacg gatcggcact
     2401 gcgtgcgcca attctgcaga caaatggcag tattcatcca caattttaaa agaaaagggg
     2461 ggattggggg gtacagtgca ggggaaagaa tagtagacat aatagcaaca gacatacaaa
     2521 ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt tcgggtttat tacagggaca
     2581 gcagagatcc agtttggtta attaaggtac cgagggccta tttcccatga ttccttcata
     2641 tttgcatata cgatacaagg ctgttagaga gataattaga attaatttga ctgtaaacac
     2701 aaagatatta gtacaaaata cgtgacgtag aaagtaataa tttcttgggt agtttgcagt
     2761 tttaaaatta tgttttaaaa tggactatca tatgcttacc gtaacttgaa agtatttcga
     2821 tttcttggct ttatatatct tgtggaaagg acgaaacacc ggattgtggt cgctcaaaac
     2881 cgttttagag ctagaaatag caagttaaaa taaggctagt ccgttatcaa cttgaaaaag
     2941 tggcaccgag tcggtgcttt tttgaattcg ctagctaggt cttgaaagga gtgggaattg
     3001 gctccggtgc ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg
     3061 gaggggtcgg caattgatcc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg
     3121 atgtcgtgta ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag
     3181 tagtcgccgt gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gaccggttct
     3241 agagcgctgc caccatggac aagaagtaca gcatcggcct ggacatcggc accaactctg
     3301 tgggctgggc cgtgatcacc gacgagtaca aggtgcccag caagaaattc aaggtgctgg
     3361 gcaacaccga ccggcacagc atcaagaaga acctgatcgg agccctgctg ttcgacagcg
     3421 gcgaaacagc cgaggccacc cggctgaaga gaaccgccag aagaagatac accagacgga
     3481 agaaccggat ctgctatctg caagagatct tcagcaacga gatggccaag gtggacgaca
     3541 gcttcttcca cagactggaa gagtccttcc tggtggaaga ggataagaag cacgagcggc
     3601 accccatctt cggcaacatc gtggacgagg tggcctacca cgagaagtac cccaccatct
     3661 accacctgag aaagaaactg gtggacagca ccgacaaggc cgacctgcgg ctgatctatc
     3721 tggccctggc ccacatgatc aagttccggg gccacttcct gatcgagggc gacctgaacc
     3781 ccgacaacag cgacgtggac aagctgttca tccagctggt gcagacctac aaccagctgt
     3841 tcgaggaaaa ccccatcaac gccagcggcg tggacgccaa ggccatcctg tctgccagac
     3901 tgagcaagag cagacggctg gaaaatctga tcgcccagct gcccggcgag aagaagaatg
     3961 gcctgttcgg aaacctgatt gccctgagcc tgggcctgac ccccaacttc aagagcaact
     4021 tcgacctggc cgaggatgcc aaactgcagc tgagcaagga cacctacgac gacgacctgg
     4081 acaacctgct ggcccagatc ggcgaccagt acgccgacct gtttctggcc gccaagaacc
     4141 tgtccgacgc catcctgctg agcgacatcc tgagagtgaa caccgagatc accaaggccc
     4201 ccctgagcgc ctctatgatc aagagatacg acgagcacca ccaggacctg accctgctga
     4261 aagctctcgt gcggcagcag ctgcctgaga agtacaaaga gattttcttc gaccagagca
     4321 agaacggcta cgccggctac attgacggcg gagccagcca ggaagagttc tacaagttca
     4381 tcaagcccat cctggaaaag atggacggca ccgaggaact gctcgtgaag ctgaacagag
     4441 aggacctgct gcggaagcag cggaccttcg acaacggcag catcccccac cagatccacc
     4501 tgggagagct gcacgccatt ctgcggcggc aggaagattt ttacccattc ctgaaggaca
     4561 accgggaaaa gatcgagaag atcctgacct tccgcatccc ctactacgtg ggccctctgg
     4621 ccaggggaaa cagcagattc gcctggatga ccagaaagag cgaggaaacc atcaccccct
     4681 ggaacttcga ggaagtggtg gacaagggcg cttccgccca gagcttcatc gagcggatga
     4741 ccaacttcga taagaacctg cccaacgaga aggtgctgcc caagcacagc ctgctgtacg
     4801 agtacttcac cgtgtataac gagctgacca aagtgaaata cgtgaccgag ggaatgagaa
     4861 agcccgcctt cctgagcggc gagcagaaaa aggccatcgt ggacctgctg ttcaagacca
     4921 accggaaagt gaccgtgaag cagctgaaag aggactactt caagaaaatc gagtgcttcg
     4981 actccgtgga aatctccggc gtggaagatc ggttcaacgc ctccctgggc acataccacg
     5041 atctgctgaa aattatcaag gacaaggact tcctggacaa tgaggaaaac gaggacattc
     5101 tggaagatat cgtgctgacc ctgacactgt ttgaggacag agagatgatc gaggaacggc
     5161 tgaaaaccta tgcccacctg ttcgacgaca aagtgatgaa gcagctgaag cggcggagat
     5221 acaccggctg gggcaggctg agccggaagc tgatcaacgg catccgggac aagcagtccg
     5281 gcaagacaat cctggatttc ctgaagtccg acggcttcgc caacagaaac ttcatgcagc
     5341 tgatccacga cgacagcctg acctttaaag aggacatcca gaaagcccag gtgtccggcc
     5401 agggcgatag cctgcacgag cacattgcca atctggccgg cagccccgcc attaagaagg
     5461 gcatcctgca gacagtgaag gtggtggacg agctcgtgaa agtgatgggc cggcacaagc
     5521 ccgagaacat cgtgatcgaa atggccagag agaaccagac cacccagaag ggacagaaga
     5581 acagccgcga gagaatgaag cggatcgaag agggcatcaa agagctgggc agccagatcc
     5641 tgaaagaaca ccccgtggaa aacacccagc tgcagaacga gaagctgtac ctgtactacc
     5701 tgcagaatgg gcgggatatg tacgtggacc aggaactgga catcaaccgg ctgtccgact
     5761 acgatgtgga ccatatcgtg cctcagagct ttctgaagga cgactccatc gacaacaagg
     5821 tgctgaccag aagcgacaag aaccggggca agagcgacaa cgtgccctcc gaagaggtcg
     5881 tgaagaagat gaagaactac tggcggcagc tgctgaacgc caagctgatt acccagagaa
     5941 agttcgacaa tctgaccaag gccgagagag gcggcctgag cgaactggat aaggccggct
     6001 tcatcaagag acagctggtg gaaacccggc agatcacaaa gcacgtggca cagatcctgg
     6061 actcccggat gaacactaag tacgacgaga atgacaagct gatccgggaa gtgaaagtga
     6121 tcaccctgaa gtccaagctg gtgtccgatt tccggaagga tttccagttt tacaaagtgc
     6181 gcgagatcaa caactaccac cacgcccacg acgcctacct gaacgccgtc gtgggaaccg
     6241 ccctgatcaa aaagtaccct aagctggaaa gcgagttcgt gtacggcgac tacaaggtgt
     6301 acgacgtgcg gaagatgatc gccaagagcg agcaggaaat cggcaaggct accgccaagt
     6361 acttcttcta cagcaacatc atgaactttt tcaagaccga gattaccctg gccaacggcg
     6421 agatccggaa gcggcctctg atcgagacaa acggcgaaac cggggagatc gtgtgggata
     6481 agggccggga ttttgccacc gtgcggaaag tgctgagcat gccccaagtg aatatcgtga
     6541 aaaagaccga ggtgcagaca ggcggcttca gcaaagagtc tatcctgccc aagaggaaca
     6601 gcgataagct gatcgccaga aagaaggact gggaccctaa gaagtacggc ggcttcgaca
     6661 gccccaccgt ggcctattct gtgctggtgg tggccaaagt ggaaaagggc aagtccaaga
     6721 aactgaagag tgtgaaagag ctgctgggga tcaccatcat ggaaagaagc agcttcgaga
     6781 agaatcccat cgactttctg gaagccaagg gctacaaaga agtgaaaaag gacctgatca
     6841 tcaagctgcc taagtactcc ctgttcgagc tggaaaacgg ccggaagaga atgctggcct
     6901 ctgccggcga actgcagaag ggaaacgaac tggccctgcc ctccaaatat gtgaacttcc
     6961 tgtacctggc cagccactat gagaagctga agggctcccc cgaggataat gagcagaaac
     7021 agctgtttgt ggaacagcac aagcactacc tggacgagat catcgagcag atcagcgagt
     7081 tctccaagag agtgatcctg gccgacgcta atctggacaa agtgctgtcc gcctacaaca
     7141 agcaccggga taagcccatc agagagcagg ccgagaatat catccacctg tttaccctga
     7201 ccaatctggg agcccctgcc gccttcaagt actttgacac caccatcgac cggaagaggt
     7261 acaccagcac caaagaggtg ctggacgcca ccctgatcca ccagagcatc accggcctgt
     7321 acgagacacg gatcgacctg tctcagctgg gaggcgacaa gcgacctgcc gccacaaaga
     7381 aggctggaca ggctaagaag aagaaagatt acaaagacga tgacgataag ggatccggcg
     7441 caacaaactt ctctctgctg aaacaagccg gagatgtcga agagaatcct ggaccgaaaa
     7501 agcctgaact caccgcgacg tctgtcgaga agtttctgat cgaaaagttc gacagcgtgt
     7561 ccgacctgat gcagctctcg gagggcgaag aatctcgtgc tttcagcttc gatgtaggag
     7621 ggcgtggata tgtcctgcgg gtaaatagct gcgccgatgg tttctacaaa gatcgttatg
     7681 tttatcggca ctttgcatcg gccgcgctcc cgattccgga agtgcttgac attggggaat
     7741 tcagcgagag cctgacctat tgcatctccc gccgtgcaca gggtgtcacg ttgcaagacc
     7801 tgcctgaaac cgaactgccc gctgttctgc agccggtcgc ggaggccatg gatgcgatcg
     7861 ctgcggccga tcttagccag acgagcgggt tcggcccatt cggaccgcaa ggaatcggtc
     7921 aatacactac atggcgtgat ttcatatgcg cgattgctga tccccatgtg tatcactggc
     7981 aaactgtgat ggacgacacc gtcagtgcgt ccgtcgcgca ggctctcgat gagctgatgc
     8041 tttgggccga ggactgcccc gaagtccggc acctcgtgca cgcggatttc ggctccaaca
     8101 atgtcctgac ggacaatggc cgcataacag cggtcattga ctggagcgag gcgatgttcg
     8161 gggattccca atacgaggtc gccaacatct tcttctggag gccgtggttg gcttgtatgg
     8221 agcagcagac gcgctacttc gagcggaggc atccggagct tgcaggatcg ccgcggctcc
     8281 gggcgtatat gctccgcatt ggtcttgacc aactctatca gagcttggtt gacggcaatt
     8341 tcgatgatgc agcttgggcg cagggtcgat gcgacgcaat cgtccgatcc ggagccggga
     8401 ctgtcgggcg tacacaaatc gcccgcagaa gcgcggccgt ctggaccgat ggctgtgtag
     8461 aagtacttcg ccgatagacg cgttaagtcg acaatcaacc tctggattac aaaatttgtg
     8521 aaagattgac tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt
     8581 taatgccttt gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata
     8641 aatcctggtt gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg
     8701 tgtgcactgt gtttgctgac gcaaccccca ctggttgggg cattgccacc acctgtcagc
     8761 tcctttccgg gactttcgct ttccccctcc ctattgccac ggcggaactc atcgccgcct
     8821 gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttgt
     8881 cggggaaatc atcgtccttt ccttggctgc tcgcctgtgt tgccacctgg attctgcgcg
     8941 ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc ggaccttcct tcccgcggcc
     9001 tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg agtcggatct
     9061 ccctttgggc cgcctccccg cgtcgacttt aagaccaatg acttacaagg cagctgtaga
     9121 tcttagccac tttttaaaag aaaagggggg actggaaggg ctaattcact cccaacgaag
     9181 acaagatctg ctttttgctt gtactgggtc tctctggtta gaccagatct gagcctggga
     9241 gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc cttgagtgct
     9301 tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc tcagaccctt
     9361 ttagtcagtg tggaaaatct ctagcagggc ccgtttaaac ccgctgatca gcctcgactg
     9421 tgccttctag ttgccagcca tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg
     9481 aaggtgccac tcccactgtc ctttcctaat aaaatgagga aattgcatcg cattgtctga
     9541 gtaggtgtca ttctattctg gggggtgggg tggggcagga cagcaagggg gaggattggg
     9601 aagacaatag caggcatgct ggggatgcgg tgggctctat ggcttctgag gcggaaagaa
     9661 ccagctgggg ctctaggggg tatccccacg cgccctgtag cggcgcatta agcgcggcgg
     9721 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     9781 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     9841 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     9901 attagggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     9961 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
    10021 ctatctcggt ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
    10081 aaaatgagct gatttaacaa aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt
    10141 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
    10201 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
    10261 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
    10321 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
    10381 agaggccgag gccgcctctg cctctgagct attccagaag tagtgaggag gcttttttgg
    10441 aggcctaggc ttttgcaaaa agctcccggg agcttgtata tccattttcg gatctgatca
    10501 gcacgtgttg acaattaatc atcggcatag tatatcggca tagtataata cgacaaggtg
    10561 aggaactaaa ccatggccaa gttgaccagt gccgttccgg tgctcaccgc gcgcgacgtc
    10621 gccggagcgg tcgagttctg gaccgaccgg ctcgggttct cccgggactt cgtggaggac
    10681 gacttcgccg gtgtggtccg ggacgacgtg accctgttca tcagcgcggt ccaggaccag
    10741 gtggtgccgg acaacaccct ggcctgggtg tgggtgcgcg gcctggacga gctgtacgcc
    10801 gagtggtcgg aggtcgtgtc cacgaacttc cgggacgcct ccgggccggc catgaccgag
    10861 atcggcgagc agccgtgggg gcgggagttc gccctgcgcg acccggccgg caactgcgtg
    10921 cacttcgtgg ccgaggagca ggactgacac gtgctacgag atttcgattc caccgccgcc
    10981 ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg ccggctggat gatcctccag
    11041 cgcggggatc tcatgctgga gttcttcgcc caccccaact tgtttattgc agcttataat
    11101 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
    11161 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgtat accgtcgacc
    11221 tctagctaga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
    11281 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
    11341 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
    11401 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
    11461 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
    11521 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
    11581 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
    11641 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
    11701 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
    11761 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
    11821 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
    11881 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
    11941 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
    12001 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
    12061 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
    12121 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
    12181 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
    12241 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
    12301 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
    12361 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
    12421 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
    12481 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
    12541 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
    12601 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
    12661 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
    12721 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
    12781 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
    12841 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
    12901 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
    12961 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
    13021 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
    13081 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
    13141 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
    13201 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
    13261 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
    13321 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
    13381 ccccgaaaag tgccacctga c
//
