LOCUS       sequence-462133-       13467 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral, CRISPR sgRNA for PCSK5.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13467)
  AUTHORS   Marohl T, Atkins KA, Wang L, Janes KA
  TITLE     PCSK5M452I is a recessive hypomorph exclusive to MCF10DCIS.com 
            cells.
  JOURNAL   Mol Cancer Res. 2025 Oct 28. doi: 10.1158/1541-7786.MCR-25-0211.
  PUBMED    41148168
REFERENCE   2  (bases 1 to 13467)
  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: "Mol Cancer 
            Res. 2025 Oct 28. doi: 10.1158/1541-7786.MCR-25-0211."
COMMENT     Sequence Label: EDCPV PCSK5_sg09
FEATURES             Location/Qualifiers
     source          1..13467
                     /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"
     misc_RNA        2878..2953
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        3015..3226
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      3240..3249
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3246..3248,3249..3569)
                     /codon_start=1
                     /product="destabilization domain that can be stabilized by 
                     Shield1 in the ProteoTuner(TM) system"
                     /label=DD
                     /note="L106P mutant of FKBP12"
                     /translation="MGVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKVDSSRDRNK
                     PFKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVEL
                     LKPE"
     CDS             3579..7682
                     /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             3579..7682
                     /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             7683..7730
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7731..7754
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             7764..7820
                     /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             join(7824..7826,7827..7829,7830..8543)
                     /codon_start=1
                     /product="yellow fluorescent protein (YFP) with fast and 
                     efficient maturation (Nagai et al., 2002)"
                     /label=Venus
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KLICTTGKLPVPWPTLVTTLGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(8130..8149)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     8477..8498
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    8559..9147
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8612..8632)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(9030..9041)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             9219..9452
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(9478..9495)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9484..9708
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      9754..10182
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9841..9860)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10051..10072
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10191..10211)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10196..10525
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      10376..10511
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10438..10457
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10573..10620
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10639..11013
                     /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    11143..11264
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11180..11199)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11234..11253
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11313..11329)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11313..11329)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11326..11348)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11337..11353
                     /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(11361..11367,11368..11385,11386..11391))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11406..11427
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11544..11561)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11715..12303)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11795..11814)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12474..13265,13266..13334))
                     /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     13097..13116
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(13335..13439)
                     /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 aacaccgcta cacgggaaag aacattggtt
     2881 ttagagctag aaatagcaag ttaaaataag gctagtccgt tatcaacttg aaaaagtggc
     2941 accgagtcgg tgcttttttg aattcgctag ctaggtcttg aaaggagtgg gaattggctc
     3001 cggtgcccgt cagtgggcag agcgcacatc gcccacagtc cccgagaagt tggggggagg
     3061 ggtcggcaat tgatccggtg cctagagaag gtggcgcggg gtaaactggg aaagtgatgt
     3121 cgtgtactgg ctccgccttt ttcccgaggg tgggggagaa ccgtatataa gtgcagtagt
     3181 cgccgtgaac gttctttttc gcaacgggtt tgccgccaga acacaggacc ggttctagag
     3241 ccaccatggg agtgcaggtg gaaaccatct ccccaggtga cgggcgcacc ttccccaagc
     3301 gcggccagac ctgtgtggtg cactacaccg ggatgcttga agatggaaag aaagtcgatt
     3361 cctcccggga cagaaacaag ccctttaagt ttatgctagg caagcaggag gtgatccgag
     3421 gctgggaaga aggggttgcc cagatgagtg tgggtcagag agccaaactg actatatctc
     3481 cagattatgc ctatggtgcc actgggcacc caggcatcat cccaccacat gccactctcg
     3541 tcttcgatgt ggagcttcta aaaccggaac cggctagcat ggacaagaag tacagcatcg
     3601 gcctggacat cggcaccaac tctgtgggct gggccgtgat caccgacgag tacaaggtgc
     3661 ccagcaagaa attcaaggtg ctgggcaaca ccgaccggca cagcatcaag aagaacctga
     3721 tcggagccct gctgttcgac agcggcgaaa cagccgaggc cacccggctg aagagaaccg
     3781 ccagaagaag atacaccaga cggaagaacc ggatctgcta tctgcaagag atcttcagca
     3841 acgagatggc caaggtggac gacagcttct tccacagact ggaagagtcc ttcctggtgg
     3901 aagaggataa gaagcacgag cggcacccca tcttcggcaa catcgtggac gaggtggcct
     3961 accacgagaa gtaccccacc atctaccacc tgagaaagaa actggtggac agcaccgaca
     4021 aggccgacct gcggctgatc tatctggccc tggcccacat gatcaagttc cggggccact
     4081 tcctgatcga gggcgacctg aaccccgaca acagcgacgt ggacaagctg ttcatccagc
     4141 tggtgcagac ctacaaccag ctgttcgagg aaaaccccat caacgccagc ggcgtggacg
     4201 ccaaggccat cctgtctgcc agactgagca agagcagacg gctggaaaat ctgatcgccc
     4261 agctgcccgg cgagaagaag aatggcctgt tcggaaacct gattgccctg agcctgggcc
     4321 tgacccccaa cttcaagagc aacttcgacc tggccgagga tgccaaactg cagctgagca
     4381 aggacaccta cgacgacgac ctggacaacc tgctggccca gatcggcgac cagtacgccg
     4441 acctgtttct ggccgccaag aacctgtccg acgccatcct gctgagcgac atcctgagag
     4501 tgaacaccga gatcaccaag gcccccctga gcgcctctat gatcaagaga tacgacgagc
     4561 accaccagga cctgaccctg ctgaaagctc tcgtgcggca gcagctgcct gagaagtaca
     4621 aagagatttt cttcgaccag agcaagaacg gctacgccgg ctacattgac ggcggagcca
     4681 gccaggaaga gttctacaag ttcatcaagc ccatcctgga aaagatggac ggcaccgagg
     4741 aactgctcgt gaagctgaac agagaggacc tgctgcggaa gcagcggacc ttcgacaacg
     4801 gcagcatccc ccaccagatc cacctgggag agctgcacgc cattctgcgg cggcaggaag
     4861 atttttaccc attcctgaag gacaaccggg aaaagatcga gaagatcctg accttccgca
     4921 tcccctacta cgtgggccct ctggccaggg gaaacagcag attcgcctgg atgaccagaa
     4981 agagcgagga aaccatcacc ccctggaact tcgaggaagt ggtggacaag ggcgcttccg
     5041 cccagagctt catcgagcgg atgaccaact tcgataagaa cctgcccaac gagaaggtgc
     5101 tgcccaagca cagcctgctg tacgagtact tcaccgtgta taacgagctg accaaagtga
     5161 aatacgtgac cgagggaatg agaaagcccg ccttcctgag cggcgagcag aaaaaggcca
     5221 tcgtggacct gctgttcaag accaaccgga aagtgaccgt gaagcagctg aaagaggact
     5281 acttcaagaa aatcgagtgc ttcgactccg tggaaatctc cggcgtggaa gatcggttca
     5341 acgcctccct gggcacatac cacgatctgc tgaaaattat caaggacaag gacttcctgg
     5401 acaatgagga aaacgaggac attctggaag atatcgtgct gaccctgaca ctgtttgagg
     5461 acagagagat gatcgaggaa cggctgaaaa cctatgccca cctgttcgac gacaaagtga
     5521 tgaagcagct gaagcggcgg agatacaccg gctggggcag gctgagccgg aagctgatca
     5581 acggcatccg ggacaagcag tccggcaaga caatcctgga tttcctgaag tccgacggct
     5641 tcgccaacag aaacttcatg cagctgatcc acgacgacag cctgaccttt aaagaggaca
     5701 tccagaaagc ccaggtgtcc ggccagggcg atagcctgca cgagcacatt gccaatctgg
     5761 ccggcagccc cgccattaag aagggcatcc tgcagacagt gaaggtggtg gacgagctcg
     5821 tgaaagtgat gggccggcac aagcccgaga acatcgtgat cgaaatggcc agagagaacc
     5881 agaccaccca gaagggacag aagaacagcc gcgagagaat gaagcggatc gaagagggca
     5941 tcaaagagct gggcagccag atcctgaaag aacaccccgt ggaaaacacc cagctgcaga
     6001 acgagaagct gtacctgtac tacctgcaga atgggcggga tatgtacgtg gaccaggaac
     6061 tggacatcaa ccggctgtcc gactacgatg tggaccatat cgtgcctcag agctttctga
     6121 aggacgactc catcgacaac aaggtgctga ccagaagcga caagaaccgg ggcaagagcg
     6181 acaacgtgcc ctccgaagag gtcgtgaaga agatgaagaa ctactggcgg cagctgctga
     6241 acgccaagct gattacccag agaaagttcg acaatctgac caaggccgag agaggcggcc
     6301 tgagcgaact ggataaggcc ggcttcatca agagacagct ggtggaaacc cggcagatca
     6361 caaagcacgt ggcacagatc ctggactccc ggatgaacac taagtacgac gagaatgaca
     6421 agctgatccg ggaagtgaaa gtgatcaccc tgaagtccaa gctggtgtcc gatttccgga
     6481 aggatttcca gttttacaaa gtgcgcgaga tcaacaacta ccaccacgcc cacgacgcct
     6541 acctgaacgc cgtcgtggga accgccctga tcaaaaagta ccctaagctg gaaagcgagt
     6601 tcgtgtacgg cgactacaag gtgtacgacg tgcggaagat gatcgccaag agcgagcagg
     6661 aaatcggcaa ggctaccgcc aagtacttct tctacagcaa catcatgaac tttttcaaga
     6721 ccgagattac cctggccaac ggcgagatcc ggaagcggcc tctgatcgag acaaacggcg
     6781 aaaccgggga gatcgtgtgg gataagggcc gggattttgc caccgtgcgg aaagtgctga
     6841 gcatgcccca agtgaatatc gtgaaaaaga ccgaggtgca gacaggcggc ttcagcaaag
     6901 agtctatcct gcccaagagg aacagcgata agctgatcgc cagaaagaag gactgggacc
     6961 ctaagaagta cggcggcttc gacagcccca ccgtggccta ttctgtgctg gtggtggcca
     7021 aagtggaaaa gggcaagtcc aagaaactga agagtgtgaa agagctgctg gggatcacca
     7081 tcatggaaag aagcagcttc gagaagaatc ccatcgactt tctggaagcc aagggctaca
     7141 aagaagtgaa aaaggacctg atcatcaagc tgcctaagta ctccctgttc gagctggaaa
     7201 acggccggaa gagaatgctg gcctctgccg gcgaactgca gaagggaaac gaactggccc
     7261 tgccctccaa atatgtgaac ttcctgtacc tggccagcca ctatgagaag ctgaagggct
     7321 cccccgagga taatgagcag aaacagctgt ttgtggaaca gcacaagcac tacctggacg
     7381 agatcatcga gcagatcagc gagttctcca agagagtgat cctggccgac gctaatctgg
     7441 acaaagtgct gtccgcctac aacaagcacc gggataagcc catcagagag caggccgaga
     7501 atatcatcca cctgtttacc ctgaccaatc tgggagcccc tgccgccttc aagtactttg
     7561 acaccaccat cgaccggaag aggtacacca gcaccaaaga ggtgctggac gccaccctga
     7621 tccaccagag catcaccggc ctgtacgaga cacggatcga cctgtctcag ctgggaggcg
     7681 acaagcgacc tgccgccaca aagaaggctg gacaggctaa gaagaagaaa gattacaaag
     7741 acgatgacga taagggatcc ggagctacta acttcagcct gctgaagcag gctggggacg
     7801 tggaggagaa ccctggacct aggatggtga gcaagggcga ggagctgttc accggggtgg
     7861 tgcccatcct ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg
     7921 agggcgaggg cgatgccacc tacggcaagc tgaccctgaa gctgatctgc accaccggca
     7981 agctgcccgt gccctggccc accctcgtga ccaccctggg ctacggcctg cagtgcttcg
     8041 cccgctaccc cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct
     8101 acgtccagga gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg
     8161 tgaagttcga gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg
     8221 aggacggcaa catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata
     8281 tcaccgccga caagcagaag aacggcatca aggccaactt caagatccgc cacaacatcg
     8341 aggacggcgg cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc
     8401 ccgtgctgct gcccgacaac cactacctga gctaccagtc cgccctgagc aaagacccca
     8461 acgagaagcg cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg
     8521 gcatggacga gctgtacaag taaggatcct aagtcgacaa tcaacctctg gattacaaaa
     8581 tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg
     8641 ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct
     8701 tgtataaatc ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg
     8761 gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct
     8821 gtcagctcct ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg
     8881 ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg
     8941 tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc ctgtgttgcc acctggattc
     9001 tgcgcgggac gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc
     9061 gcggcctgct gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc
     9121 ggatctccct ttgggccgcc tccccgcgtc gactttaaga ccaatgactt acaaggcagc
     9181 tgtagatctt agccactttt taaaagaaaa ggggggactg gaagggctaa ttcactccca
     9241 acgaagacaa gatctgcttt ttgcttgtac tgggtctctc tggttagacc agatctgagc
     9301 ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa gcttgccttg
     9361 agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga gatccctcag
     9421 acccttttag tcagtgtgga aaatctctag cagggcccgt ttaaacccgc tgatcagcct
     9481 cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga
     9541 ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt
     9601 gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg
     9661 attgggaaga caatagcagg catgctgggg atgcggtggg ctctatggct tctgaggcgg
     9721 aaagaaccag ctggggctct agggggtatc cccacgcgcc ctgtagcggc gcattaagcg
     9781 cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg
     9841 ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc
     9901 taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa
     9961 aacttgatta gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc
    10021 ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac
    10081 tcaaccctat ctcggtctat tcttttgatt tataagggat tttgccgatt tcggcctatt
    10141 ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg
    10201 tcagttaggg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
    10261 tctcaattag tcagcaacca ggtgtggaaa gtccccaggc tccccagcag gcagaagtat
    10321 gcaaagcatg catctcaatt agtcagcaac catagtcccg cccctaactc cgcccatccc
    10381 gcccctaact ccgcccagtt ccgcccattc tccgccccat ggctgactaa ttttttttat
    10441 ttatgcagag gccgaggccg cctctgcctc tgagctattc cagaagtagt gaggaggctt
    10501 ttttggaggc ctaggctttt gcaaaaagct cccgggagct tgtatatcca ttttcggatc
    10561 tgatcagcac gtgttgacaa ttaatcatcg gcatagtata tcggcatagt ataatacgac
    10621 aaggtgagga actaaaccat ggccaagttg accagtgccg ttccggtgct caccgcgcgc
    10681 gacgtcgccg gagcggtcga gttctggacc gaccggctcg ggttctcccg ggacttcgtg
    10741 gaggacgact tcgccggtgt ggtccgggac gacgtgaccc tgttcatcag cgcggtccag
    10801 gaccaggtgg tgccggacaa caccctggcc tgggtgtggg tgcgcggcct ggacgagctg
    10861 tacgccgagt ggtcggaggt cgtgtccacg aacttccggg acgcctccgg gccggccatg
    10921 accgagatcg gcgagcagcc gtgggggcgg gagttcgccc tgcgcgaccc ggccggcaac
    10981 tgcgtgcact tcgtggccga ggagcaggac tgacacgtgc tacgagattt cgattccacc
    11041 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc
    11101 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct
    11161 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
    11221 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg
    11281 tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt
    11341 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt
    11401 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg
    11461 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
    11521 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
    11581 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
    11641 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
    11701 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
    11761 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
    11821 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
    11881 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
    11941 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
    12001 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
    12061 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
    12121 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg
    12181 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
    12241 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
    12301 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
    12361 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
    12421 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
    12481 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
    12541 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
    12601 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
    12661 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
    12721 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
    12781 gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
    12841 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
    12901 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
    12961 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
    13021 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
    13081 ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
    13141 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
    13201 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
    13261 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
    13321 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
    13381 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
    13441 acatttcccc gaaaagtgcc acctgac
//
