LOCUS       sequence-482849-       11741 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral vector expressing Cas9-P2A-Blasticidin and two sgRNAs 
            targeting RPP21.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11741)
  AUTHORS   Che R, Panah M, Mirani B, Knowles K, Ostapovich A, Majumdar D, Chen 
            X, DeSimone J, White W, Noonan M, Luo H, Alexandrov A
  TITLE     Identification of human pathways acting on nuclear non-coding RNAs 
            using the Mirror forward genetic approach.
  JOURNAL   Nat Commun. 2025 May 22;16(1):4741. doi: 10.1038/s41467-025-59998-3.
  PUBMED    40399278
REFERENCE   2  (bases 1 to 11741)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2025-05-22"; volume: "16(1):4741. doi"; pages: " 
            10.1038/s41467-025-59998-3"
COMMENT     Sequence Label: LentiCRISPR-B_2xsgRNAs_RPP21 (pAVA3259)
FEATURES             Location/Qualifiers
     source          1..11741
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        5..231
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             232..412
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    459..584
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1077..1310
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1495..1539
                     /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"
     promoter        1717..1965
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1717..1737
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     1888..1907
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        1987..2062
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_RNA        2339..2414
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_feature    2462..2579
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2648..2859
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      2879..2888
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2888..2911
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2918..2938
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2963..7063
                     /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             2963..7063
                     /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             7064..7111
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7127..7183
                     /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             7184..7582
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    7598..8186
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7651..7671)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8069..8080)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             8258..8491
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    8569..8690
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8606..8625)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8660..8679
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      8730..8865
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(8885..8904)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(8886..8904)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(8914..8931)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(8914..8930)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8923..8945)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      9072..9527
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9159..9178)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9369..9390
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        9553..9657
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(9658..9726,9727..10518)
                     /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(9876..9895)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      10689..11277
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     11178..11197
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     11431..11448
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    11565..11586
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(11601..11606,11607..11624,11625..11631)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11639..11655
                     /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     11644..11666
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     11663..11679
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     11663..11679
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     11698..11718
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        11700..11718
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 ttaatgtagt cttatgcaat actcttgtag tcttgcaaca tggtaacgat gagttagcaa
       61 catgccttac aaggagagaa aaagcaccgt gcatgccgat tggtggaagt aaggtggtac
      121 gatcgtgcct tattaggaag gcaacagacg ggtctgacat ggattggacg aaccactgaa
      181 ttgccgcatt gcagagatat tgtatttaag tgcctagctc gatacataaa cgggtctctc
      241 tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag
      301 cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct
      361 ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag cagtggcgcc
      421 cgaacaggga cttgaaagcg aaagggaaac cagaggagct ctctcgacgc aggactcggc
      481 ttgctgaagc gcgcacggca agaggcgagg ggcggcgact ggtgagtacg ccaaaaattt
      541 tgactagcgg aggctagaag gagagagatg ggtgcgagag cgtcagtatt aagcggggga
      601 gaattagatc gcgatgggaa aaaattcggt taaggccagg gggaaagaaa aaatataaat
      661 taaaacatat agtatgggca agcagggagc tagaacgatt cgcagttaat cctggcctgt
      721 tagaaacatc agaaggctgt agacaaatac tgggacagct acaaccatcc cttcagacag
      781 gatcagaaga acttagatca ttatataata cagtagcaac cctctattgt gtgcatcaaa
      841 ggatagagat aaaagacacc aaggaagctt tagacaagat agaggaagag caaaacaaaa
      901 gtaagaccac cgcacagcaa gcggccgctg atcttcagac ctggaggagg agatatgagg
      961 gacaattgga gaagtgaatt atataaatat aaagtagtaa aaattgaacc attaggagta
     1021 gcacccacca aggcaaagag aagagtggtg cagagagaaa aaagagcagt gggaatagga
     1081 gctttgttcc ttgggttctt gggagcagca ggaagcacta tgggcgcagc gtcaatgacg
     1141 ctgacggtac aggccagaca attattgtct ggtatagtgc agcagcagaa caatttgctg
     1201 agggctattg aggcgcaaca gcatctgttg caactcacag tctggggcat caagcagctc
     1261 caggcaagaa tcctggctgt ggaaagatac ctaaaggatc aacagctcct ggggatttgg
     1321 ggttgctctg gaaaactcat ttgcaccact gctgtgcctt ggaatgctag ttggagtaat
     1381 aaatctctgg aacagatttg gaatcacacg acctggatgg agtgggacag agaaattaac
     1441 aattacacaa gcttaataca ctccttaatt gaagaatcgc aaaaccagca agaaaagaat
     1501 gaacaagaat tattggaatt agataaatgg gcaagtttgt ggaattggtt taacataaca
     1561 aattggctgt ggtatataaa attattcata atgatagtag gaggcttggt aggtttaaga
     1621 atagtttttg ctgtactttc tatagtgaat agagttaggc agggatattc accattatcg
     1681 tttcagaccc acctcccaac cccgagggga cccagagagg gcctatttcc catgattcct
     1741 tcatatttgc atatacgata caaggctgtt agagagataa ttagaattaa tttgactgta
     1801 aacacaaaga tattagtaca aaatacgtga cgtagaaagt aataatttct tgggtagttt
     1861 gcagttttaa aattatgttt taaaatggac tatcatatgc ttaccgtaac ttgaaagtat
     1921 ttcgatttct tggctttata tatcttgtgg aaaggacgaa acaccggagg ctcaacttcc
     1981 tgtaccgttt tagagctaga aatagcaagt taaaataagg ctagtccgtt atcaacttga
     2041 aaaagtggca ccgagtcggt gcttttttga attcctgcag tatttagcat gccccaccca
     2101 tctgcaaggc attctggata gtgtcaaaac agccggaaat caagtccgtt tatctcaaac
     2161 tttagcattt tgggaataaa tgatatttgc tatgctggtt aaattagatt ttagttaaat
     2221 ttcctgctga agctctagta cgataagcaa cttgacctaa gtgtaaagtt gagacttcct
     2281 tcaggtttat atagcttgtg cgccgcttgg gtacctcgcg aagcggctcg tcttgcgggt
     2341 tttagagcta gaaatagcaa gttaaaataa ggctagtccg ttatcaactt gaaaaagtgg
     2401 caccgagtcg gtgctttttt gaattctaga tcttgagaca aatggcagta ttcatccaca
     2461 attttaaaag aaaagggggg attggggggt acagtgcagg ggaaagaata gtagacataa
     2521 tagcaacaga catacaaact aaagaattac aaaaacaaat tacaaaaatt caaaattttc
     2581 gggtttatta cagggacagc agagatccac tttggcgccg gctcgagtgg ctccggtgcc
     2641 cgtcagtggg cagagcgcac atcgcccaca gtccccgaga agttgggggg aggggtcggc
     2701 aattgaaccg gtgcctagag aaggtggcgc ggggtaaact gggaaagtga tgtcgtgtac
     2761 tggctccgcc tttttcccga gggtggggga gaaccgtata taagtgcagt agtcgccgtg
     2821 aacgttcttt ttcgcaacgg gtttgccgcc agaacacagg tgtcgtgacg cgggatccgc
     2881 caccatggat tacaaagacg atgacgataa gatggcccca aagaagaagc ggaaggtcgg
     2941 tatccacgga gtcccagcag ccgacaagaa gtacagcatc ggcctggaca tcggcaccaa
     3001 ctctgtgggc tgggccgtga tcaccgacga gtacaaggtg cccagcaaga aattcaaggt
     3061 gctgggcaac accgaccggc acagcatcaa gaagaacctg atcggagccc tgctgttcga
     3121 cagcggcgaa acagccgagg ccacccggct gaagagaacc gccagaagaa gatacaccag
     3181 acggaagaac cggatctgct atctgcaaga gatcttcagc aacgagatgg ccaaggtgga
     3241 cgacagcttc ttccacagac tggaagagtc cttcctggtg gaagaggata agaagcacga
     3301 gcggcacccc atcttcggca acatcgtgga cgaggtggcc taccacgaga agtaccccac
     3361 catctaccac ctgagaaaga aactggtgga cagcaccgac aaggccgacc tgcggctgat
     3421 ctatctggcc ctggcccaca tgatcaagtt ccggggccac ttcctgatcg agggcgacct
     3481 gaaccccgac aacagcgacg tggacaagct gttcatccag ctggtgcaga cctacaacca
     3541 gctgttcgag gaaaacccca tcaacgccag cggcgtggac gccaaggcca tcctgtctgc
     3601 cagactgagc aagagcagac ggctggaaaa tctgatcgcc cagctgcccg gcgagaagaa
     3661 gaatggcctg ttcggaaacc tgattgccct gagcctgggc ctgaccccca acttcaagag
     3721 caacttcgac ctggccgagg atgccaaact gcagctgagc aaggacacct acgacgacga
     3781 cctggacaac ctgctggccc agatcggcga ccagtacgcc gacctgtttc tggccgccaa
     3841 gaacctgtcc gacgccatcc tgctgagcga catcctgaga gtgaacaccg agatcaccaa
     3901 ggcccccctg agcgcctcta tgatcaagag atacgacgag caccaccagg acctgaccct
     3961 gctgaaagct ctcgtgcggc agcagctgcc tgagaagtac aaagagattt tcttcgacca
     4021 gagcaagaac ggctacgccg gctacattga cggcggagcc agccaggaag agttctacaa
     4081 gttcatcaag cccatcctgg aaaagatgga cggcaccgag gaactgctcg tgaagctgaa
     4141 cagagaggac ctgctgcgga agcagcggac cttcgacaac ggcagcatcc cccaccagat
     4201 ccacctggga gagctgcacg ccattctgcg gcggcaggaa gatttttacc cattcctgaa
     4261 ggacaaccgg gaaaagatcg agaagatcct gaccttccgc atcccctact acgtgggccc
     4321 tctggccagg ggaaacagca gattcgcctg gatgaccaga aagagcgagg aaaccatcac
     4381 cccctggaac ttcgaggaag tggtggacaa gggcgcttcc gcccagagct tcatcgagcg
     4441 gatgaccaac ttcgataaga acctgcccaa cgagaaggtg ctgcccaagc acagcctgct
     4501 gtacgagtac ttcaccgtgt ataacgagct gaccaaagtg aaatacgtga ccgagggaat
     4561 gagaaagccc gccttcctga gcggcgagca gaaaaaggcc atcgtggacc tgctgttcaa
     4621 gaccaaccgg aaagtgaccg tgaagcagct gaaagaggac tacttcaaga aaatcgagtg
     4681 cttcgactcc gtggaaatct ccggcgtgga agatcggttc aacgcctccc tgggcacata
     4741 ccacgatctg ctgaaaatta tcaaggacaa ggacttcctg gacaatgagg aaaacgagga
     4801 cattctggaa gatatcgtgc tgaccctgac actgtttgag gacagagaga tgatcgagga
     4861 acggctgaaa acctatgccc acctgttcga cgacaaagtg atgaagcagc tgaagcggcg
     4921 gagatacacc ggctggggca ggctgagccg gaagctgatc aacggcatcc gggacaagca
     4981 gtccggcaag acaatcctgg atttcctgaa gtccgacggc ttcgccaaca gaaacttcat
     5041 gcagctgatc cacgacgaca gcctgacctt taaagaggac atccagaaag cccaggtgtc
     5101 cggccagggc gatagcctgc acgagcacat tgccaatctg gccggcagcc ccgccattaa
     5161 gaagggcatc ctgcagacag tgaaggtggt ggacgagctc gtgaaagtga tgggccggca
     5221 caagcccgag aacatcgtga tcgaaatggc cagagagaac cagaccaccc agaagggaca
     5281 gaagaacagc cgcgagagaa tgaagcggat cgaagagggc atcaaagagc tgggcagcca
     5341 gatcctgaaa gaacaccccg tggaaaacac ccagctgcag aacgagaagc tgtacctgta
     5401 ctacctgcag aatgggcggg atatgtacgt ggaccaggaa ctggacatca accggctgtc
     5461 cgactacgat gtggaccata tcgtgcctca gagctttctg aaggacgact ccatcgacaa
     5521 caaggtgctg accagaagcg acaagaaccg gggcaagagc gacaacgtgc cctccgaaga
     5581 ggtcgtgaag aagatgaaga actactggcg gcagctgctg aacgccaagc tgattaccca
     5641 gagaaagttc gacaatctga ccaaggccga gagaggcggc ctgagcgaac tggataaggc
     5701 cggcttcatc aagagacagc tggtggaaac ccggcagatc acaaagcacg tggcacagat
     5761 cctggactcc cggatgaaca ctaagtacga cgagaatgac aagctgatcc gggaagtgaa
     5821 agtgatcacc ctgaagtcca agctggtgtc cgatttccgg aaggatttcc agttttacaa
     5881 agtgcgcgag atcaacaact accaccacgc ccacgacgcc tacctgaacg ccgtcgtggg
     5941 aaccgccctg atcaaaaagt accctaagct ggaaagcgag ttcgtgtacg gcgactacaa
     6001 ggtgtacgac gtgcggaaga tgatcgccaa gagcgagcag gaaatcggca aggctaccgc
     6061 caagtacttc ttctacagca acatcatgaa ctttttcaag accgagatta ccctggccaa
     6121 cggcgagatc cggaagcggc ctctgatcga gacaaacggc gaaaccgggg agatcgtgtg
     6181 ggataagggc cgggattttg ccaccgtgcg gaaagtgctg agcatgcccc aagtgaatat
     6241 cgtgaaaaag accgaggtgc agacaggcgg cttcagcaaa gagtctatcc tgcccaagag
     6301 gaacagcgat aagctgatcg ccagaaagaa ggactgggac cctaagaagt acggcggctt
     6361 cgacagcccc accgtggcct attctgtgct ggtggtggcc aaagtggaaa agggcaagtc
     6421 caagaaactg aagagtgtga aagagctgct ggggatcacc atcatggaaa gaagcagctt
     6481 cgagaagaat cccatcgact ttctggaagc caagggctac aaagaagtga aaaaggacct
     6541 gatcatcaag ctgcctaagt actccctgtt cgagctggaa aacggccgga agagaatgct
     6601 ggcctctgcc ggcgaactgc agaagggaaa cgaactggcc ctgccctcca aatatgtgaa
     6661 cttcctgtac ctggccagcc actatgagaa gctgaagggc tcccccgagg ataatgagca
     6721 gaaacagctg tttgtggaac agcacaagca ctacctggac gagatcatcg agcagatcag
     6781 cgagttctcc aagagagtga tcctggccga cgctaatctg gacaaagtgc tgtccgccta
     6841 caacaagcac cgggataagc ccatcagaga gcaggccgag aatatcatcc acctgtttac
     6901 cctgaccaat ctgggagccc ctgccgcctt caagtacttt gacaccacca tcgaccggaa
     6961 gaggtacacc agcaccaaag aggtgctgga cgccaccctg atccaccaga gcatcaccgg
     7021 cctgtacgag acacggatcg acctgtctca gctgggaggc gacaagcgtc ctgctgctac
     7081 taagaaagct ggtcaagcta agaaaaagaa agctagcggc agcggcgcca ccaacttcag
     7141 cctgctgaag caggccggcg acgtggagga gaaccccggc cccatggcca agcctttgtc
     7201 tcaagaagaa tccaccctca ttgaaagagc aacggctaca atcaacagca tccccatctc
     7261 tgaagactac agcgtcgcca gcgcagctct ctctagcgac ggccgcatct tcactggtgt
     7321 caatgtatat cattttactg ggggaccttg tgcagaactc gtggtgctgg gcactgctgc
     7381 tgctgcggca gctggcaacc tgacttgtat cgtcgcgatc ggaaatgaga acaggggcat
     7441 cttgagcccc tgcggacggt gccgacaggt gcttctcgat ctgcatcctg ggatcaaagc
     7501 catagtgaag gacagtgatg gacagccgac ggcagttggg attcgtgaat tgctgccctc
     7561 tggttatgtg tgggagggct aaacgcgtta agtcgacaat caacctctgg attacaaaat
     7621 ttgtgaaaga ttgactggta ttcttaacta tgttgctcct tttacgctat gtggatacgc
     7681 tgctttaatg cctttgtatc atgctattgc ttcccgtatg gctttcattt tctcctcctt
     7741 gtataaatcc tggttgctgt ctctttatga ggagttgtgg cccgttgtca ggcaacgtgg
     7801 cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt tggggcattg ccaccacctg
     7861 tcagctcctt tccgggactt tcgctttccc cctccctatt gccacggcgg aactcatcgc
     7921 cgcctgcctt gcccgctgct ggacaggggc tcggctgttg ggcactgaca attccgtggt
     7981 gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc tgtgttgcca cctggattct
     8041 gcgcgggacg tccttctgct acgtcccttc ggccctcaat ccagcggacc ttccttcccg
     8101 cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc cttcgccctc agacgagtcg
     8161 gatctccctt tgggccgcct ccccgcgtcg actttaagac caatgactta caaggcagct
     8221 gtagatctta gccacttttt aaaagaaaag gggggactgg aagggctaat tcactcccaa
     8281 cgaagacaag atctgctttt tgcttgtact gggtctctct ggttagacca gatctgagcc
     8341 tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag cttgccttga
     8401 gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag atccctcaga
     8461 cccttttagt cagtgtggaa aatctctagc agtacgtata gtagttcatg tcatcttatt
     8521 attcagtatt tataacttgc aaagaaatga atatcagaga gtgagaggaa cttgtttatt
     8581 gcagcttata atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt
     8641 ttttcactgc attctagttg tggtttgtcc aaactcatca atgtatctta tcatgtctgg
     8701 ctctagctat cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     8761 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctcgg
     8821 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggg acgtacccaa
     8881 ttcgccctat agtgagtcgt attacgcgcg ctcactggcc gtcgttttac aacgtcgtga
     8941 ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag
     9001 ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa
     9061 tggcgaatgg gacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     9121 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     9181 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
     9241 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg gtgatggttc
     9301 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
     9361 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cggtctattc
     9421 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
     9481 acaaaaattt aacgcgaatt ttaacaaaat attaacgctt acaatttagg tggcactttt
     9541 cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat
     9601 ccgctcatga gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg
     9661 agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt
     9721 tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga
     9781 gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa
     9841 gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt
     9901 attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt
     9961 gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc
    10021 agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga
    10081 ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat
    10141 cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct
    10201 gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc
    10261 cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg
    10321 gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc
    10381 ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg
    10441 acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca
    10501 ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta gattgattta
    10561 aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc
    10621 aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa
    10681 ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca
    10741 ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta
    10801 actggcttca gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc
    10861 caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca
    10921 gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta
    10981 ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag
    11041 cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt
    11101 cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc
    11161 acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac
    11221 ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac
    11281 gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc
    11341 tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat
    11401 accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag
    11461 cgcccaatac gcaaaccgcc tctccccgcg cgttggccga ttcattaatg cagctggcac
    11521 gacaggtttc ccgactggaa agcgggcagt gagcgcaacg caattaatgt gagttagctc
    11581 actcattagg caccccaggc tttacacttt atgcttccgg ctcgtatgtt gtgtggaatt
    11641 gtgagcggat aacaatttca cacaggaaac agctatgacc atgattacgc caagcgcgca
    11701 attaaccctc actaaaggga acaaaagctg gagctgcaag c
//
