LOCUS       sequence-479164-       11389 bp    DNA     circular SYN 04-AUG-2025
DEFINITION  Lentiviral vector expressing Cas9-P2A-Blasticidin and an sgRNA 
            targeting RPP20.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11389)
  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 11389)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Aug 4, 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_sgRNA_RPP20 (pAVA3546)
FEATURES             Location/Qualifiers
     source          1..11389
                     /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_feature    2110..2227
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2296..2507
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      2527..2536
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2536..2559
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             2566..2586
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2611..6711
                     /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             2611..6711
                     /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             6712..6759
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             6775..6831
                     /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             6832..7230
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    7246..7834
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7299..7319)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(7717..7728)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             7906..8139
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    8217..8338
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8254..8273)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8308..8327
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      8378..8513
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(8533..8552)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(8534..8552)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(8562..8579)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(8562..8578)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8571..8593)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      8720..9175
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8807..8826)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9017..9038
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        9201..9305
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(9306..9374,9375..10166)
                     /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(9524..9543)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      10337..10925
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     10826..10845
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     11079..11096
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    11213..11234
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(11249..11254,11255..11272,11273..11279)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11287..11303
                     /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     11292..11314
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     11311..11327
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     11311..11327
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     11346..11366
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        11348..11366
                     /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 acaccgggct gccaatacct
     1981 ccaccggttt tagagctaga aatagcaagt taaaataagg ctagtccgtt atcaacttga
     2041 aaaagtggca ccgagtcggt gcttttttga attctagatc ttgagacaaa tggcagtatt
     2101 catccacaat tttaaaagaa aaggggggat tggggggtac agtgcagggg aaagaatagt
     2161 agacataata gcaacagaca tacaaactaa agaattacaa aaacaaatta caaaaattca
     2221 aaattttcgg gtttattaca gggacagcag agatccactt tggcgccggc tcgagtggct
     2281 ccggtgcccg tcagtgggca gagcgcacat cgcccacagt ccccgagaag ttggggggag
     2341 gggtcggcaa ttgaaccggt gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg
     2401 tcgtgtactg gctccgcctt tttcccgagg gtgggggaga accgtatata agtgcagtag
     2461 tcgccgtgaa cgttcttttt cgcaacgggt ttgccgccag aacacaggtg tcgtgacgcg
     2521 ggatccgcca ccatggatta caaagacgat gacgataaga tggccccaaa gaagaagcgg
     2581 aaggtcggta tccacggagt cccagcagcc gacaagaagt acagcatcgg cctggacatc
     2641 ggcaccaact ctgtgggctg ggccgtgatc accgacgagt acaaggtgcc cagcaagaaa
     2701 ttcaaggtgc tgggcaacac cgaccggcac agcatcaaga agaacctgat cggagccctg
     2761 ctgttcgaca gcggcgaaac agccgaggcc acccggctga agagaaccgc cagaagaaga
     2821 tacaccagac ggaagaaccg gatctgctat ctgcaagaga tcttcagcaa cgagatggcc
     2881 aaggtggacg acagcttctt ccacagactg gaagagtcct tcctggtgga agaggataag
     2941 aagcacgagc ggcaccccat cttcggcaac atcgtggacg aggtggccta ccacgagaag
     3001 taccccacca tctaccacct gagaaagaaa ctggtggaca gcaccgacaa ggccgacctg
     3061 cggctgatct atctggccct ggcccacatg atcaagttcc ggggccactt cctgatcgag
     3121 ggcgacctga accccgacaa cagcgacgtg gacaagctgt tcatccagct ggtgcagacc
     3181 tacaaccagc tgttcgagga aaaccccatc aacgccagcg gcgtggacgc caaggccatc
     3241 ctgtctgcca gactgagcaa gagcagacgg ctggaaaatc tgatcgccca gctgcccggc
     3301 gagaagaaga atggcctgtt cggaaacctg attgccctga gcctgggcct gacccccaac
     3361 ttcaagagca acttcgacct ggccgaggat gccaaactgc agctgagcaa ggacacctac
     3421 gacgacgacc tggacaacct gctggcccag atcggcgacc agtacgccga cctgtttctg
     3481 gccgccaaga acctgtccga cgccatcctg ctgagcgaca tcctgagagt gaacaccgag
     3541 atcaccaagg cccccctgag cgcctctatg atcaagagat acgacgagca ccaccaggac
     3601 ctgaccctgc tgaaagctct cgtgcggcag cagctgcctg agaagtacaa agagattttc
     3661 ttcgaccaga gcaagaacgg ctacgccggc tacattgacg gcggagccag ccaggaagag
     3721 ttctacaagt tcatcaagcc catcctggaa aagatggacg gcaccgagga actgctcgtg
     3781 aagctgaaca gagaggacct gctgcggaag cagcggacct tcgacaacgg cagcatcccc
     3841 caccagatcc acctgggaga gctgcacgcc attctgcggc ggcaggaaga tttttaccca
     3901 ttcctgaagg acaaccggga aaagatcgag aagatcctga ccttccgcat cccctactac
     3961 gtgggccctc tggccagggg aaacagcaga ttcgcctgga tgaccagaaa gagcgaggaa
     4021 accatcaccc cctggaactt cgaggaagtg gtggacaagg gcgcttccgc ccagagcttc
     4081 atcgagcgga tgaccaactt cgataagaac ctgcccaacg agaaggtgct gcccaagcac
     4141 agcctgctgt acgagtactt caccgtgtat aacgagctga ccaaagtgaa atacgtgacc
     4201 gagggaatga gaaagcccgc cttcctgagc ggcgagcaga aaaaggccat cgtggacctg
     4261 ctgttcaaga ccaaccggaa agtgaccgtg aagcagctga aagaggacta cttcaagaaa
     4321 atcgagtgct tcgactccgt ggaaatctcc ggcgtggaag atcggttcaa cgcctccctg
     4381 ggcacatacc acgatctgct gaaaattatc aaggacaagg acttcctgga caatgaggaa
     4441 aacgaggaca ttctggaaga tatcgtgctg accctgacac tgtttgagga cagagagatg
     4501 atcgaggaac ggctgaaaac ctatgcccac ctgttcgacg acaaagtgat gaagcagctg
     4561 aagcggcgga gatacaccgg ctggggcagg ctgagccgga agctgatcaa cggcatccgg
     4621 gacaagcagt ccggcaagac aatcctggat ttcctgaagt ccgacggctt cgccaacaga
     4681 aacttcatgc agctgatcca cgacgacagc ctgaccttta aagaggacat ccagaaagcc
     4741 caggtgtccg gccagggcga tagcctgcac gagcacattg ccaatctggc cggcagcccc
     4801 gccattaaga agggcatcct gcagacagtg aaggtggtgg acgagctcgt gaaagtgatg
     4861 ggccggcaca agcccgagaa catcgtgatc gaaatggcca gagagaacca gaccacccag
     4921 aagggacaga agaacagccg cgagagaatg aagcggatcg aagagggcat caaagagctg
     4981 ggcagccaga tcctgaaaga acaccccgtg gaaaacaccc agctgcagaa cgagaagctg
     5041 tacctgtact acctgcagaa tgggcgggat atgtacgtgg accaggaact ggacatcaac
     5101 cggctgtccg actacgatgt ggaccatatc gtgcctcaga gctttctgaa ggacgactcc
     5161 atcgacaaca aggtgctgac cagaagcgac aagaaccggg gcaagagcga caacgtgccc
     5221 tccgaagagg tcgtgaagaa gatgaagaac tactggcggc agctgctgaa cgccaagctg
     5281 attacccaga gaaagttcga caatctgacc aaggccgaga gaggcggcct gagcgaactg
     5341 gataaggccg gcttcatcaa gagacagctg gtggaaaccc ggcagatcac aaagcacgtg
     5401 gcacagatcc tggactcccg gatgaacact aagtacgacg agaatgacaa gctgatccgg
     5461 gaagtgaaag tgatcaccct gaagtccaag ctggtgtccg atttccggaa ggatttccag
     5521 ttttacaaag tgcgcgagat caacaactac caccacgccc acgacgccta cctgaacgcc
     5581 gtcgtgggaa ccgccctgat caaaaagtac cctaagctgg aaagcgagtt cgtgtacggc
     5641 gactacaagg tgtacgacgt gcggaagatg atcgccaaga gcgagcagga aatcggcaag
     5701 gctaccgcca agtacttctt ctacagcaac atcatgaact ttttcaagac cgagattacc
     5761 ctggccaacg gcgagatccg gaagcggcct ctgatcgaga caaacggcga aaccggggag
     5821 atcgtgtggg ataagggccg ggattttgcc accgtgcgga aagtgctgag catgccccaa
     5881 gtgaatatcg tgaaaaagac cgaggtgcag acaggcggct tcagcaaaga gtctatcctg
     5941 cccaagagga acagcgataa gctgatcgcc agaaagaagg actgggaccc taagaagtac
     6001 ggcggcttcg acagccccac cgtggcctat tctgtgctgg tggtggccaa agtggaaaag
     6061 ggcaagtcca agaaactgaa gagtgtgaaa gagctgctgg ggatcaccat catggaaaga
     6121 agcagcttcg agaagaatcc catcgacttt ctggaagcca agggctacaa agaagtgaaa
     6181 aaggacctga tcatcaagct gcctaagtac tccctgttcg agctggaaaa cggccggaag
     6241 agaatgctgg cctctgccgg cgaactgcag aagggaaacg aactggccct gccctccaaa
     6301 tatgtgaact tcctgtacct ggccagccac tatgagaagc tgaagggctc ccccgaggat
     6361 aatgagcaga aacagctgtt tgtggaacag cacaagcact acctggacga gatcatcgag
     6421 cagatcagcg agttctccaa gagagtgatc ctggccgacg ctaatctgga caaagtgctg
     6481 tccgcctaca acaagcaccg ggataagccc atcagagagc aggccgagaa tatcatccac
     6541 ctgtttaccc tgaccaatct gggagcccct gccgccttca agtactttga caccaccatc
     6601 gaccggaaga ggtacaccag caccaaagag gtgctggacg ccaccctgat ccaccagagc
     6661 atcaccggcc tgtacgagac acggatcgac ctgtctcagc tgggaggcga caagcgtcct
     6721 gctgctacta agaaagctgg tcaagctaag aaaaagaaag ctagcggcag cggcgccacc
     6781 aacttcagcc tgctgaagca ggccggcgac gtggaggaga accccggccc catggccaag
     6841 cctttgtctc aagaagaatc caccctcatt gaaagagcaa cggctacaat caacagcatc
     6901 cccatctctg aagactacag cgtcgccagc gcagctctct ctagcgacgg ccgcatcttc
     6961 actggtgtca atgtatatca ttttactggg ggaccttgtg cagaactcgt ggtgctgggc
     7021 actgctgctg ctgcggcagc tggcaacctg acttgtatcg tcgcgatcgg aaatgagaac
     7081 aggggcatct tgagcccctg cggacggtgc cgacaggtgc ttctcgatct gcatcctggg
     7141 atcaaagcca tagtgaagga cagtgatgga cagccgacgg cagttgggat tcgtgaattg
     7201 ctgccctctg gttatgtgtg ggagggctaa acgcgttaag tcgacaatca acctctggat
     7261 tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt
     7321 ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc
     7381 tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg
     7441 caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc
     7501 accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa
     7561 ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat
     7621 tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc tgctcgcctg tgttgccacc
     7681 tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt
     7741 ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag
     7801 acgagtcgga tctccctttg ggccgcctcc ccgcgtcgac tttaagacca atgacttaca
     7861 aggcagctgt agatcttagc cactttttaa aagaaaaggg gggactggaa gggctaattc
     7921 actcccaacg aagacaagat ctgctttttg cttgtactgg gtctctctgg ttagaccaga
     7981 tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct
     8041 tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat
     8101 ccctcagacc cttttagtca gtgtggaaaa tctctagcag tacgtatagt agttcatgtc
     8161 atcttattat tcagtattta taacttgcaa agaaatgaat atcagagagt gagaggaact
     8221 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     8281 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     8341 atgtctggct ctagctatcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     8401 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     8461 cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctagggac
     8521 gtacccaatt cgccctatag tgagtcgtat tacgcgcgct cactggccgt cgttttacaa
     8581 cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct
     8641 ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc
     8701 agcctgaatg gcgaatggga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     8761 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
     8821 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     8881 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
     8941 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     9001 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
     9061 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
     9121 ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttaggtg
     9181 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     9241 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     9301 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     9361 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     9421 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     9481 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     9541 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     9601 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     9661 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     9721 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     9781 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     9841 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     9901 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     9961 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
    10021 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
    10081 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
    10141 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
    10201 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
    10261 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
    10321 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
    10381 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
    10441 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
    10501 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
    10561 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
    10621 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
    10681 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
    10741 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
    10801 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
    10861 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
    10921 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
    10981 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt
    11041 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag
    11101 cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
    11161 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga
    11221 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt
    11281 gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca
    11341 agcgcgcaat taaccctcac taaagggaac aaaagctgga gctgcaagc
//
