LOCUS       sequence-450313-       13791 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of CRISPRi PELO sgRNA #2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13791)
  AUTHORS   Borck PC, Boyle I, Jankovic K, Bick N, Foster K, Lau AC, 
            Parker-Burns LI, Lubicki DA, Li T, Borah AA, Lofaso NJ, Das Sharma 
            S, Chan T, Kishen RV, Adeagbo A, Raghavan S, Aquilanti E, Prensner 
            JR, Krill-Burger JM, Golub TR, Campbell CD, Dempster JM, Chan EM, 
            Vazquez F
  TITLE     SKI complex loss renders 9p21.3-deleted or MSI-H cancers dependent 
            on PELO.
  JOURNAL   Nature. 2025 Feb 5. doi: 10.1038/s41586-024-08509-3.
  PUBMED    39910293
REFERENCE   2  (bases 1 to 13791)
  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: "Nature. 
            2025 Feb 5. doi: 10.1038/s41586-024-08509-3."
COMMENT     Sequence Label: pXPR_BRD051d_sgCiPELO #2
FEATURES             Location/Qualifiers
     source          1..13791
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(47..66)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        238..617
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..817
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     772..792
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             835..1015
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1062..1187
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1680..1913
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2098..2142
                     /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    2440..2557
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2608..2856
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2608..2628
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2779..2798
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     promoter        3025..3236
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     protein_bind    3262..3286
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             3316..3501
                     /codon_start=1
                     /product="Kruppel-associated box (KRAB) transcriptional 
                     repression domain from the human zinc finger protein ZNF10 
                     (Margolin et al., 1994)"
                     /label=KRAB
                     /translation="RTLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGY
                     QLTKPDVILRLEKGEEP"
     CDS             join(3523..3549,3550..3552,3553..6039,6040..6042,
                     6043..7626)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the H840A mutation in the HNH 
                     catalytic domain"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             3523..7626
                     /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="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             7630..7656
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             7636..7662
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     CDS             7675..7695
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7702..7722
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7765..7821
                     /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             7831..8856
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVHRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     misc_feature    8878..9466
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8931..8951)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(9349..9360)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             9544..9777
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(9802..9819)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9808..10032
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      10078..10506
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10165..10184)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10375..10396
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10515..10535)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10520..10849
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      10700..10835
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10762..10781
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10897..10944
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10963..11337
                     /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    11467..11588
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11504..11523)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11558..11577
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11637..11653)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11637..11653)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11650..11672)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11661..11677
                     /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(11685..11691,11692..11709,11710..11715))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11730..11751
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11868..11885)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(12039..12627)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(12119..12138)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12798..13589,13590..13658))
                     /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     13421..13440
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(13659..13763)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
      241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
      301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct
      841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
      901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
      961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
     1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
     1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
     1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
     1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
     1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
     1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
     1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
     1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca
     1501 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg
     1561 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga
     1621 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata
     1681 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg
     1741 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg
     1801 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag
     1861 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt
     1921 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt
     1981 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt
     2041 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag
     2101 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata
     2161 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta
     2221 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta
     2281 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa
     2341 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt
     2401 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat
     2461 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa
     2521 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag
     2581 agatccagtt tggttaatta aggtaccgag ggcctatttc ccatgattcc ttcatatttg
     2641 catatacgat acaaggctgt tagagagata attagaatta atttgactgt aaacacaaag
     2701 atattagtac aaaatacgtg acgtagaaag taataatttc ttgggtagtt tgcagtttta
     2761 aaattatgtt ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta tttcgatttc
     2821 ttggctttat atatcttgtg gaaaggacga aacaccgtgc cagcgggaac tgtgtaggtt
     2881 tcagagctac agcagaaatg ctgtagcaag ttgaaataag gctagtccgt tatcaacttg
     2941 aaaaagtggc accgagtcgg tgcttttttg aattcgctag ctaggtcttg aaaggagtgg
     3001 gaattggctc cggtgcccgt cagtgggcag agcgcacatc gcccacagtc cccgagaagt
     3061 tggggggagg ggtcggcaat tgatccggtg cctagagaag gtggcgcggg gtaaactggg
     3121 aaagtgatgt cgtgtactgg ctccgccttt ttcccgaggg tgggggagaa ccgtatataa
     3181 gtgcagtagt cgccgtgaac gttctttttc gcaacgggtt tgccgccaga acacaggacc
     3241 ggttctagca tcgattgatc aacaagtttg tacaaaaaag ttggcatgga tgctaagtca
     3301 ctaactgcct ggtcccggac actggtgacc ttcaaggatg tatttgtgga cttcaccagg
     3361 gaggagtgga agctgctgga cactgctcag cagatcgtgt acagaaatgt gatgctggag
     3421 aactataaga acctggtttc cttgggttat cagcttacta agccagatgt gatcctccgg
     3481 ttggagaagg gagaagagcc cggcggttcc ggcggagggt cgatggacaa gaagtacagc
     3541 atcggcctgg ccatcggcac caactctgtg ggctgggccg tgatcaccga cgagtacaag
     3601 gtgcccagca agaaattcaa ggtgctgggc aacaccgacc ggcacagcat caagaagaac
     3661 ctgatcggcg ccctgctgtt cgacagcgga gaaacagccg aggccacccg gctgaagaga
     3721 accgccagaa gaagatacac cagacggaag aaccggatct gctatctgca agagatcttc
     3781 agcaacgaga tggccaaggt ggacgacagc ttcttccaca gactggaaga gtccttcctg
     3841 gtggaagagg ataagaagca cgagcggcac cccatcttcg gcaacatcgt ggacgaggtg
     3901 gcctaccacg agaagtaccc caccatctac cacctgagaa agaaactggt ggacagcacc
     3961 gacaaggccg acctgcggct gatctatctg gccctggccc acatgatcaa gttccggggc
     4021 cacttcctga tcgagggcga cctgaacccc gacaacagcg acgtggacaa gctgttcatc
     4081 cagctggtgc agacctacaa ccagctgttc gaggaaaacc ccatcaacgc cagcggcgtg
     4141 gacgccaagg ccatcctgtc tgccagactg agcaagagca gacggctgga aaatctgatc
     4201 gcccagctgc ccggcgagaa gaagaatggc ctgttcggca acctgattgc cctgagcctg
     4261 ggcctgaccc ccaacttcaa gagcaacttc gacctggccg aggatgccaa actgcagctg
     4321 agcaaggaca cctacgacga cgacctggac aacctgctgg cccagatcgg cgaccagtac
     4381 gccgacctgt ttctggccgc caagaacctg tccgacgcca tcctgctgag cgacatcctg
     4441 agagtgaaca ccgagatcac caaggccccc ctgagcgcct ctatgatcaa gagatacgac
     4501 gagcaccacc aggacctgac cctgctgaaa gctctcgtgc ggcagcagct gcctgagaag
     4561 tacaaagaga ttttcttcga ccagagcaag aacggctacg ccggctacat cgatggcgga
     4621 gccagccagg aagagttcta caagttcatc aagcccatcc tggaaaagat ggacggcacc
     4681 gaggaactgc tcgtgaagct gaacagagag gacctgctgc ggaagcagcg gaccttcgac
     4741 aacggcagca tcccccacca gatccacctg ggagagctgc acgccattct gcggcggcag
     4801 gaagattttt acccattcct gaaggacaac cgggaaaaga tcgagaagat cctgaccttc
     4861 cgcatcccct actacgtggg ccctctggcc aggggaaaca gcagattcgc ctggatgacc
     4921 agaaagagcg aggaaaccat caccccctgg aacttcgagg aagtggtgga caagggcgcc
     4981 agcgcccaga gcttcatcga gcggatgacc aacttcgata agaacctgcc caacgagaag
     5041 gtgctgccca agcacagcct gctgtacgag tacttcaccg tgtacaacga gctgaccaaa
     5101 gtgaaatacg tgaccgaggg aatgagaaag cccgccttcc tgagcggcga gcagaaaaaa
     5161 gccatcgtgg acctgctgtt caagaccaac cggaaagtga ccgtgaagca gctgaaagag
     5221 gactacttca agaaaatcga gtgcttcgac tccgtggaaa tctccggcgt ggaagatcgg
     5281 ttcaacgcct ccctgggcac ataccacgat ctgctgaaaa ttatcaagga caaggacttc
     5341 ctggacaatg aggaaaacga ggacattctg gaagatatcg tgctgaccct gacactgttt
     5401 gaggacagag agatgatcga ggaacggctg aaaacctatg cccacctgtt cgacgacaaa
     5461 gtgatgaagc agctgaagcg gcggagatac accggctggg gcaggctgag ccggaagctg
     5521 atcaacggca tccgggacaa gcagtccggc aagacaatcc tggatttcct gaagtccgac
     5581 ggcttcgcca acagaaactt catgcagctg atccacgacg acagcctgac ctttaaagag
     5641 gacatccaga aagcccaggt gtccggccag ggcgatagcc tgcacgagca cattgccaat
     5701 ctggccggca gccccgccat taagaagggc atcctgcaga cagtgaaggt ggtggacgag
     5761 ctcgtgaaag tgatgggccg gcacaagccc gagaacatcg tgatcgaaat ggccagagag
     5821 aaccagacca cccagaaggg acagaagaac agccgcgaga gaatgaagcg gatcgaagag
     5881 ggcatcaaag agctgggcag ccagatcctg aaagaacacc ccgtggaaaa cacccagctg
     5941 cagaacgaga agctgtacct gtactacctg cagaatgggc gggatatgta cgtggaccag
     6001 gaactggaca tcaaccggct gtccgactac gatgtggacg ctatcgtgcc tcagagcttt
     6061 ctgaaggacg actccatcga taacaaagtg ctgactcgga gcgacaagaa ccggggcaag
     6121 agcgacaacg tgccctccga agaggtcgtg aagaagatga agaactactg gcgccagctg
     6181 ctgaatgcca agctgattac ccagaggaag ttcgacaatc tgaccaaggc cgagagaggc
     6241 ggcctgagcg aactggataa ggccggcttc atcaagagac agctggtgga aacccggcag
     6301 atcacaaagc acgtggcaca gatcctggac tcccggatga acactaagta cgacgagaac
     6361 gacaaactga tccgggaagt gaaagtgatc accctgaagt ccaagctggt gtccgatttc
     6421 cggaaggatt tccagtttta caaagtgcgc gagatcaaca actaccacca cgcccacgac
     6481 gcctacctga acgccgtcgt gggaaccgcc ctgatcaaaa agtaccctaa gctggaaagc
     6541 gagttcgtgt acggcgacta caaggtgtac gacgtgcgga agatgatcgc caagagcgag
     6601 caggaaatcg gcaaggctac cgccaagtac ttcttctaca gcaacatcat gaactttttc
     6661 aagaccgaga ttaccctggc caacggcgag atccggaagc ggcctctgat cgagacaaac
     6721 ggcgaaacag gcgagatcgt gtgggataag ggccgggact ttgccaccgt gcggaaagtg
     6781 ctgtctatgc cccaagtgaa tatcgtgaaa aagaccgagg tgcagacagg cggcttcagc
     6841 aaagagtcta tcctgcccaa gaggaacagc gacaagctga tcgccagaaa gaaggactgg
     6901 gaccctaaga agtacggcgg cttcgacagc cccaccgtgg cctattctgt gctggtggtg
     6961 gccaaagtgg aaaagggcaa gtccaagaaa ctgaagagtg tgaaagagct gctggggatc
     7021 accatcatgg aaagaagcag cttcgagaag aatcccatcg actttctgga agccaagggc
     7081 tacaaagaag tgaaaaagga cctgatcatc aagctgccta agtactccct gttcgagctg
     7141 gaaaacggcc ggaagagaat gctggcctct gccggcgaac tgcagaaggg aaacgaactg
     7201 gccctgccct ccaaatatgt gaacttcctg tacctggcca gccactatga gaagctgaag
     7261 ggctcccccg aggataatga gcagaaacag ctgtttgtgg aacagcacaa acactacctg
     7321 gacgagatca tcgagcagat cagcgagttc tccaagagag tgatcctggc cgacgctaat
     7381 ctggacaagg tgctgagcgc ctacaacaag cacagagaca agcctatcag agagcaggcc
     7441 gagaatatca tccacctgtt taccctgacc aatctgggag cccctgccgc cttcaagtac
     7501 tttgacacca ccatcgaccg gaagaggtac accagcacca aagaggtgct ggacgccacc
     7561 ctgatccacc agagcatcac cggcctgtac gagacacgga tcgacctgtc tcagctggga
     7621 ggcgacgcct atccctatga cgtgcccgat tatgccagcc tgggcagcgg ctcccccaag
     7681 aaaaaacgca aggtggaaga tcctaagaaa aagcggaaag tggacggcat tggtagtggg
     7741 agcaacggca gcagcggatc cggcgcaaca aacttctctc tgctgaaaca agccggagat
     7801 gtcgaagaga atcctggacc gggcggatcg atgaaaaagc ctgaactcac cgcgacgtct
     7861 gtcgagaagt ttctgatcga aaagttcgac agcgtgtccg acctgatgca gctctcggag
     7921 ggcgaagaat ctcgtgcttt cagcttcgat gtaggagggc gtggatatgt cctgcgggta
     7981 aatagctgcg ccgatggttt ctacaaagat cgttatgttc atcggcactt tgcatcggcc
     8041 gcgctcccga ttccggaagt gcttgacatt ggggaattta gcgagagcct gacctattgc
     8101 atctcccgcc gtgcacaggg tgtcacgttg caagacctgc ctgaaaccga actgcccgct
     8161 gttctgcagc cggtcgcgga ggccatggat gcgatcgctg cggccgatct tagccagacg
     8221 agcgggttcg gcccattcgg accgcaagga atcggtcaat acactacgtg gcgtgatttc
     8281 atatgcgcga ttgctgatcc ccatgtgtat cactggcaaa ctgtgatgga cgacaccgtc
     8341 agtgcgtccg tcgcgcaggc tctcgatgag ctgatgcttt gggccgagga ctgccccgaa
     8401 gtccggcacc tcgtgcacgc ggatttcggc tccaacaatg tcctgacgga caatggccgc
     8461 ataacagcgg tcattgactg gagcgaggcg atgttcgggg attcccaata cgaggtcgcc
     8521 aacatcttct tctggaggcc gtggttggct tgtatggagc agcagacgcg ctacttcgag
     8581 cggaggcatc cggagcttgc aggatcgccg cggctccggg cgtatatgct ccgcattggt
     8641 cttgaccaac tctatcagag cttggttgac ggcaatttcg atgatgcagc ttgggcgcag
     8701 ggtcgatgcg acgcaatcgt ccgatccgga gccgggactg tcgggcgtac acaaatcgcc
     8761 cgcagaagcg cggccgtctg gaccgatggc tgtgtagaag tactcgccga tagtggaaac
     8821 cgacgcccca gcactcgtcc gagggcaaag gaatgagtta acacgcgtta agtcgacaat
     8881 caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct
     8941 tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg
     9001 gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg
     9061 cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt
     9121 tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt
     9181 gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg
     9241 ggcactgaca attccgtggt gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc
     9301 tgtgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat
     9361 ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc
     9421 cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcctcg aggtcgactt
     9481 taagaccaat gacttacaag gcagctgtag atcttagcca ctttttaaaa gaaaaggggg
     9541 gactggaagg gctaattcac tcccaacgaa gacaagatct gctttttgct tgtactgggt
     9601 ctctctggtt agaccagatc tgagcctggg agctctctgg ctaactaggg aacccactgc
     9661 ttaagcctca ataaagcttg ccttgagtgc ttcaagtagt gtgtgcccgt ctgttgtgtg
     9721 actctggtaa ctagagatcc ctcagaccct tttagtcagt gtggaaaatc tctagcaggg
     9781 ccgtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt
     9841 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     9901 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     9961 tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg
    10021 tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg
    10081 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
    10141 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
    10201 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
    10261 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
    10321 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
    10381 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
    10441 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
    10501 cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
    10561 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
    10621 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
    10681 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
    10741 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct
    10801 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg
    10861 agcttgtata tccattttcg gatctgatca gcacgtgttg acaattaatc atcggcatag
    10921 tatatcggca tagtataata cgacaaggtg aggaactaaa ccatggccaa gttgaccagt
    10981 gccgttccgg tgctcaccgc gcgcgacgtc gccggagcgg tcgagttctg gaccgaccgg
    11041 ctcgggttct cccgggactt cgtggaggac gacttcgccg gtgtggtccg ggacgacgtg
    11101 accctgttca tcagcgcggt ccaggaccag gtggtgccgg acaacaccct ggcctgggtg
    11161 tgggtgcgcg gcctggacga gctgtacgcc gagtggtcgg aggtcgtgtc cacgaacttc
    11221 cgggacgcct ccgggccggc catgaccgag atcggcgagc agccgtgggg gcgggagttc
    11281 gccctgcgcg acccggccgg caactgcgtg cacttcgtgg ccgaggagca ggactgacac
    11341 gtgctacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt
    11401 ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc
    11461 caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
    11521 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
    11581 gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca
    11641 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
    11701 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
    11761 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
    11821 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
    11881 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
    11941 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
    12001 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
    12061 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
    12121 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
    12181 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
    12241 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
    12301 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
    12361 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
    12421 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
    12481 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
    12541 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
    12601 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
    12661 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
    12721 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
    12781 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
    12841 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
    12901 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
    12961 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
    13021 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
    13081 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
    13141 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
    13201 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
    13261 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
    13321 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
    13381 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
    13441 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
    13501 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
    13561 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
    13621 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
    13681 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
    13741 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga c
//
