LOCUS       sequence-394136-       12824 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Cas9 and sgRNA control guide targeting intergenic region.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12824)
  AUTHORS   So J, Mabe NW, Englinger B, Chow KH, Moyer SM, Yerrum S, Trissal MC,
            Marques JG, Kwon JJ, Shim B, Pal S, Panditharatna E, Quinn T, 
            Schaefer DA, Jeong D, Mayhew DL, Hwang J, Beroukhim R, Ligon KL, 
            Stegmaier K, Filbin MG, Hahn WC
  TITLE     VRK1 as a synthetic lethal target in VRK2 promoter-methylated 
            cancers of the nervous system.
  JOURNAL   JCI Insight. 2022 Oct 10;7(19):e158755. doi: 
            10.1172/jci.insight.158755.
  PUBMED    36040810
REFERENCE   2  (bases 1 to 12824)
  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: "JCI 
            Insight."; date: "2022-10-10"; volume: "7(19):e158755. doi"; pages: 
            " 10.1172/jci.insight.158755"
COMMENT     Sequence Label: LentiCas9-sgCh2-2-Blast
FEATURES             Location/Qualifiers
     source          1..12824
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(202..219)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    208..432
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      478..906
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(565..584)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     775..796
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(915..935)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        920..1249
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1100..1235
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1162..1181
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        1297..1344
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             1363..1737
                     /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    1867..1988
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1904..1923)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1958..1977
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2037..2053)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2037..2053)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2050..2072)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2061..2077
                     /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(2085..2091,2092..2109,2110..2115))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2130..2151
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2268..2285)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2439..3027)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2519..2538)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3198..3989,3990..4058))
                     /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     3821..3840
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4059..4163)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4238..4257)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        4429..4808
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4810..5008
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4963..4983
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             5026..5206
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5253..5378
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    5871..6104
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6289..6333
                     /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    6631..6748
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        6799..7047
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     6799..6819
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     6970..6989
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        7069..7144
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        7206..7417
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      7436..7445
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             7442..11545
                     /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             7442..11545
                     /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             11546..11593
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             11594..11617
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             11627..11683
                     /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             11684..12082
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     misc_feature    12107..12695
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(12160..12180)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(12578..12589)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             join(12767..12824,1..176)
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
ORIGIN
        1 tctctggtta gaccagatct gagcctggga gctctctggc taactaggga acccactgct
       61 taagcctcaa taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga
      121 ctctggtaac tagagatccc tcagaccctt ttagtcagtg tggaaaatct ctagcagggc
      181 ccgtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt
      241 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
      301 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
      361 tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg
      421 tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg
      481 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
      541 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
      601 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
      661 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
      721 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
      781 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
      841 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
      901 cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
      961 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     1021 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     1081 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     1141 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct
     1201 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg
     1261 agcttgtata tccattttcg gatctgatca gcacgtgttg acaattaatc atcggcatag
     1321 tatatcggca tagtataata cgacaaggtg aggaactaaa ccatggccaa gttgaccagt
     1381 gccgttccgg tgctcaccgc gcgcgacgtc gccggagcgg tcgagttctg gaccgaccgg
     1441 ctcgggttct cccgggactt cgtggaggac gacttcgccg gtgtggtccg ggacgacgtg
     1501 accctgttca tcagcgcggt ccaggaccag gtggtgccgg acaacaccct ggcctgggtg
     1561 tgggtgcgcg gcctggacga gctgtacgcc gagtggtcgg aggtcgtgtc cacgaacttc
     1621 cgggacgcct ccgggccggc catgaccgag atcggcgagc agccgtgggg gcgggagttc
     1681 gccctgcgcg acccggccgg caactgcgtg cacttcgtgg ccgaggagca ggactgacac
     1741 gtgctacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt
     1801 ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc
     1861 caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     1921 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     1981 gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca
     2041 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     2101 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     2161 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
     2221 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     2281 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     2341 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     2401 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     2461 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     2521 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     2581 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     2641 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     2701 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     2761 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     2821 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     2881 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     2941 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     3001 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     3061 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     3121 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     3181 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     3241 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     3301 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     3361 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     3421 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     3481 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     3541 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     3601 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     3661 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     3721 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     3781 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     3841 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     3901 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     3961 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     4021 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     4081 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     4141 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga
     4201 tcgggagatc tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat
     4261 agttaagcca gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca
     4321 aaatttaagc tacaacaagg caaggcttga ccgacaattg catgaagaat ctgcttaggg
     4381 ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgcgttga cattgattat
     4441 tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt
     4501 tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc
     4561 cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac
     4621 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
     4681 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc
     4741 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
     4801 ttaccatggt gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac
     4861 ggggatttcc aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc
     4921 aacgggactt tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc
     4981 gtgtacggtg ggaggtctat ataagcagcg cgttttgcct gtactgggtc tctctggtta
     5041 gaccagatct gagcctggga gctctctggc taactaggga acccactgct taagcctcaa
     5101 taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac
     5161 tagagatccc tcagaccctt ttagtcagtg tggaaaatct ctagcagtgg cgcccgaaca
     5221 gggacttgaa agcgaaaggg aaaccagagg agctctctcg acgcaggact cggcttgctg
     5281 aagcgcgcac ggcaagaggc gaggggcggc gactggtgag tacgccaaaa attttgacta
     5341 gcggaggcta gaaggagaga gatgggtgcg agagcgtcag tattaagcgg gggagaatta
     5401 gatcgcgatg ggaaaaaatt cggttaaggc cagggggaaa gaaaaaatat aaattaaaac
     5461 atatagtatg ggcaagcagg gagctagaac gattcgcagt taatcctggc ctgttagaaa
     5521 catcagaagg ctgtagacaa atactgggac agctacaacc atcccttcag acaggatcag
     5581 aagaacttag atcattatat aatacagtag caaccctcta ttgtgtgcat caaaggatag
     5641 agataaaaga caccaaggaa gctttagaca agatagagga agagcaaaac aaaagtaaga
     5701 ccaccgcaca gcaagcggcc gctgatcttc agacctggag gaggagatat gagggacaat
     5761 tggagaagtg aattatataa atataaagta gtaaaaattg aaccattagg agtagcaccc
     5821 accaaggcaa agagaagagt ggtgcagaga gaaaaaagag cagtgggaat aggagctttg
     5881 ttccttgggt tcttgggagc agcaggaagc actatgggcg cagcgtcaat gacgctgacg
     5941 gtacaggcca gacaattatt gtctggtata gtgcagcagc agaacaattt gctgagggct
     6001 attgaggcgc aacagcatct gttgcaactc acagtctggg gcatcaagca gctccaggca
     6061 agaatcctgg ctgtggaaag atacctaaag gatcaacagc tcctggggat ttggggttgc
     6121 tctggaaaac tcatttgcac cactgctgtg ccttggaatg ctagttggag taataaatct
     6181 ctggaacaga tttggaatca cacgacctgg atggagtggg acagagaaat taacaattac
     6241 acaagcttaa tacactcctt aattgaagaa tcgcaaaacc agcaagaaaa gaatgaacaa
     6301 gaattattgg aattagataa atgggcaagt ttgtggaatt ggtttaacat aacaaattgg
     6361 ctgtggtata taaaattatt cataatgata gtaggaggct tggtaggttt aagaatagtt
     6421 tttgctgtac tttctatagt gaatagagtt aggcagggat attcaccatt atcgtttcag
     6481 acccacctcc caaccccgag gggacccgac aggcccgaag gaatagaaga agaaggtgga
     6541 gagagagaca gagacagatc cattcgatta gtgaacggat cggcactgcg tgcgccaatt
     6601 ctgcagacaa atggcagtat tcatccacaa ttttaaaaga aaagggggga ttggggggta
     6661 cagtgcaggg gaaagaatag tagacataat agcaacagac atacaaacta aagaattaca
     6721 aaaacaaatt acaaaaattc aaaattttcg ggtttattac agggacagca gagatccagt
     6781 ttggttaatt aaggtaccga gggcctattt cccatgattc cttcatattt gcatatacga
     6841 tacaaggctg ttagagagat aattagaatt aatttgactg taaacacaaa gatattagta
     6901 caaaatacgt gacgtagaaa gtaataattt cttgggtagt ttgcagtttt aaaattatgt
     6961 tttaaaatgg actatcatat gcttaccgta acttgaaagt atttcgattt cttggcttta
     7021 tatatcttgt ggaaaggacg aaacaccggg tgtgcgtatg aagcagtggt tttagagcta
     7081 gaaatagcaa gttaaaataa ggctagtccg ttatcaactt gaaaaagtgg caccgagtcg
     7141 gtgctttttt gaattcgcta gctaggtctt gaaaggagtg ggaattggct ccggtgcccg
     7201 tcagtgggca gagcgcacat cgcccacagt ccccgagaag ttggggggag gggtcggcaa
     7261 ttgatccggt gcctagagaa ggtggcgcgg ggtaaactgg gaaagtgatg tcgtgtactg
     7321 gctccgcctt tttcccgagg gtgggggaga accgtatata agtgcagtag tcgccgtgaa
     7381 cgttcttttt cgcaacgggt ttgccgccag aacacaggac cggttctaga gcgctgccac
     7441 catggacaag aagtacagca tcggcctgga catcggcacc aactctgtgg gctgggccgt
     7501 gatcaccgac gagtacaagg tgcccagcaa gaaattcaag gtgctgggca acaccgaccg
     7561 gcacagcatc aagaagaacc tgatcggagc cctgctgttc gacagcggcg aaacagccga
     7621 ggccacccgg ctgaagagaa ccgccagaag aagatacacc agacggaaga accggatctg
     7681 ctatctgcaa gagatcttca gcaacgagat ggccaaggtg gacgacagct tcttccacag
     7741 actggaagag tccttcctgg tggaagagga taagaagcac gagcggcacc ccatcttcgg
     7801 caacatcgtg gacgaggtgg cctaccacga gaagtacccc accatctacc acctgagaaa
     7861 gaaactggtg gacagcaccg acaaggccga cctgcggctg atctatctgg ccctggccca
     7921 catgatcaag ttccggggcc acttcctgat cgagggcgac ctgaaccccg acaacagcga
     7981 cgtggacaag ctgttcatcc agctggtgca gacctacaac cagctgttcg aggaaaaccc
     8041 catcaacgcc agcggcgtgg acgccaaggc catcctgtct gccagactga gcaagagcag
     8101 acggctggaa aatctgatcg cccagctgcc cggcgagaag aagaatggcc tgttcggaaa
     8161 cctgattgcc ctgagcctgg gcctgacccc caacttcaag agcaacttcg acctggccga
     8221 ggatgccaaa ctgcagctga gcaaggacac ctacgacgac gacctggaca acctgctggc
     8281 ccagatcggc gaccagtacg ccgacctgtt tctggccgcc aagaacctgt ccgacgccat
     8341 cctgctgagc gacatcctga gagtgaacac cgagatcacc aaggcccccc tgagcgcctc
     8401 tatgatcaag agatacgacg agcaccacca ggacctgacc ctgctgaaag ctctcgtgcg
     8461 gcagcagctg cctgagaagt acaaagagat tttcttcgac cagagcaaga acggctacgc
     8521 cggctacatt gacggcggag ccagccagga agagttctac aagttcatca agcccatcct
     8581 ggaaaagatg gacggcaccg aggaactgct cgtgaagctg aacagagagg acctgctgcg
     8641 gaagcagcgg accttcgaca acggcagcat cccccaccag atccacctgg gagagctgca
     8701 cgccattctg cggcggcagg aagattttta cccattcctg aaggacaacc gggaaaagat
     8761 cgagaagatc ctgaccttcc gcatccccta ctacgtgggc cctctggcca ggggaaacag
     8821 cagattcgcc tggatgacca gaaagagcga ggaaaccatc accccctgga acttcgagga
     8881 agtggtggac aagggcgctt ccgcccagag cttcatcgag cggatgacca acttcgataa
     8941 gaacctgccc aacgagaagg tgctgcccaa gcacagcctg ctgtacgagt acttcaccgt
     9001 gtataacgag ctgaccaaag tgaaatacgt gaccgaggga atgagaaagc ccgccttcct
     9061 gagcggcgag cagaaaaagg ccatcgtgga cctgctgttc aagaccaacc ggaaagtgac
     9121 cgtgaagcag ctgaaagagg actacttcaa gaaaatcgag tgcttcgact ccgtggaaat
     9181 ctccggcgtg gaagatcggt tcaacgcctc cctgggcaca taccacgatc tgctgaaaat
     9241 tatcaaggac aaggacttcc tggacaatga ggaaaacgag gacattctgg aagatatcgt
     9301 gctgaccctg acactgtttg aggacagaga gatgatcgag gaacggctga aaacctatgc
     9361 ccacctgttc gacgacaaag tgatgaagca gctgaagcgg cggagataca ccggctgggg
     9421 caggctgagc cggaagctga tcaacggcat ccgggacaag cagtccggca agacaatcct
     9481 ggatttcctg aagtccgacg gcttcgccaa cagaaacttc atgcagctga tccacgacga
     9541 cagcctgacc tttaaagagg acatccagaa agcccaggtg tccggccagg gcgatagcct
     9601 gcacgagcac attgccaatc tggccggcag ccccgccatt aagaagggca tcctgcagac
     9661 agtgaaggtg gtggacgagc tcgtgaaagt gatgggccgg cacaagcccg agaacatcgt
     9721 gatcgaaatg gccagagaga accagaccac ccagaaggga cagaagaaca gccgcgagag
     9781 aatgaagcgg atcgaagagg gcatcaaaga gctgggcagc cagatcctga aagaacaccc
     9841 cgtggaaaac acccagctgc agaacgagaa gctgtacctg tactacctgc agaatgggcg
     9901 ggatatgtac gtggaccagg aactggacat caaccggctg tccgactacg atgtggacca
     9961 tatcgtgcct cagagctttc tgaaggacga ctccatcgac aacaaggtgc tgaccagaag
    10021 cgacaagaac cggggcaaga gcgacaacgt gccctccgaa gaggtcgtga agaagatgaa
    10081 gaactactgg cggcagctgc tgaacgccaa gctgattacc cagagaaagt tcgacaatct
    10141 gaccaaggcc gagagaggcg gcctgagcga actggataag gccggcttca tcaagagaca
    10201 gctggtggaa acccggcaga tcacaaagca cgtggcacag atcctggact cccggatgaa
    10261 cactaagtac gacgagaatg acaagctgat ccgggaagtg aaagtgatca ccctgaagtc
    10321 caagctggtg tccgatttcc ggaaggattt ccagttttac aaagtgcgcg agatcaacaa
    10381 ctaccaccac gcccacgacg cctacctgaa cgccgtcgtg ggaaccgccc tgatcaaaaa
    10441 gtaccctaag ctggaaagcg agttcgtgta cggcgactac aaggtgtacg acgtgcggaa
    10501 gatgatcgcc aagagcgagc aggaaatcgg caaggctacc gccaagtact tcttctacag
    10561 caacatcatg aactttttca agaccgagat taccctggcc aacggcgaga tccggaagcg
    10621 gcctctgatc gagacaaacg gcgaaaccgg ggagatcgtg tgggataagg gccgggattt
    10681 tgccaccgtg cggaaagtgc tgagcatgcc ccaagtgaat atcgtgaaaa agaccgaggt
    10741 gcagacaggc ggcttcagca aagagtctat cctgcccaag aggaacagcg ataagctgat
    10801 cgccagaaag aaggactggg accctaagaa gtacggcggc ttcgacagcc ccaccgtggc
    10861 ctattctgtg ctggtggtgg ccaaagtgga aaagggcaag tccaagaaac tgaagagtgt
    10921 gaaagagctg ctggggatca ccatcatgga aagaagcagc ttcgagaaga atcccatcga
    10981 ctttctggaa gccaagggct acaaagaagt gaaaaaggac ctgatcatca agctgcctaa
    11041 gtactccctg ttcgagctgg aaaacggccg gaagagaatg ctggcctctg ccggcgaact
    11101 gcagaaggga aacgaactgg ccctgccctc caaatatgtg aacttcctgt acctggccag
    11161 ccactatgag aagctgaagg gctcccccga ggataatgag cagaaacagc tgtttgtgga
    11221 acagcacaag cactacctgg acgagatcat cgagcagatc agcgagttct ccaagagagt
    11281 gatcctggcc gacgctaatc tggacaaagt gctgtccgcc tacaacaagc accgggataa
    11341 gcccatcaga gagcaggccg agaatatcat ccacctgttt accctgacca atctgggagc
    11401 ccctgccgcc ttcaagtact ttgacaccac catcgaccgg aagaggtaca ccagcaccaa
    11461 agaggtgctg gacgccaccc tgatccacca gagcatcacc ggcctgtacg agacacggat
    11521 cgacctgtct cagctgggag gcgacaagcg acctgccgcc acaaagaagg ctggacaggc
    11581 taagaagaag aaagattaca aagacgatga cgataaggga tccggcgcaa caaacttctc
    11641 tctgctgaaa caagccggag atgtcgaaga gaatcctgga ccgatggcca agcctttgtc
    11701 tcaagaagaa tccaccctca ttgaaagagc aacggctaca atcaacagca tccccatctc
    11761 tgaagactac agcgtcgcca gcgcagctct ctctagcgac ggccgcatct tcactggtgt
    11821 caatgtatat cattttactg ggggaccttg tgcagaactc gtggtgctgg gcactgctgc
    11881 tgctgcggca gctggcaacc tgacttgtat cgtcgcgatc ggaaatgaga acaggggcat
    11941 cttgagcccc tgcggacggt gccgacaggt gcttctcgat ctgcatcctg ggatcaaagc
    12001 catagtgaag gacagtgatg gacagccgac ggcagttggg attcgtgaat tgctgccctc
    12061 tggttatgtg tgggagggct aagaattcga tatcaagctt atcggtaatc aacctctgga
    12121 ttacaaaatt tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg
    12181 tggatacgct gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt
    12241 ctcctccttg tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag
    12301 gcaacgtggc gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc
    12361 caccacctgt cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga
    12421 actcatcgcc gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa
    12481 ttccgtggtg ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct gtgttgccac
    12541 ctggattctg cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct
    12601 tccttcccgc ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca
    12661 gacgagtcgg atctcccttt gggccgcctc cccgcgtcga ctttaagacc aatgacttac
    12721 aaggcagctg tagatcttag ccacttttta aaagaaaagg ggggactgga agggctaatt
    12781 cactcccaac gaagacaaga tctgcttttt gcttgtactg ggtc
//
