LOCUS       sequence-385833-       11590 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral CRISPR-Cas9 plasmid containing gRNA targeting exon 1 of 
            human Tollip. Used for generation of Tollip protein knockouts in 
            human cell lines..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11590)
  AUTHORS   Ryan TA, Phillips EO, Collier CL, Jb Robinson A, Routledge D, Wood 
            RE, Assar EA, Tumbarello DA
  TITLE     Tollip coordinates Parkin-dependent trafficking of 
            mitochondrial-derived vesicles.
  JOURNAL   EMBO J. 2020 Jun 2;39(11):e102539. doi: 10.15252/embj.2019102539. 
            Epub 2020 Apr 20.
  PUBMED    32311122
REFERENCE   2  (bases 1 to 11590)
  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: "EMBO J."; 
            date: "2020-06-2"; volume: "39(11):e102539. doi"; pages: " 
            10.15252/embj.2019102539. Epub 2020 Apr 20"
COMMENT     Sequence Label: pLentiCRISPRv1 sgTollip
FEATURES             Location/Qualifiers
     source          1..11590
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     133..152
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     386..403
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    520..541
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(556..561,562..579,580..586)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    594..610
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     599..621
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     618..634
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     618..634
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     653..673
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        655..673
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        701..927
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             928..1108
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1155..1280
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1773..2006
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2191..2235
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     promoter        2413..2661
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2413..2433
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2584..2603
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2683..2758
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_feature    2806..2923
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2992..3203
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      3223..3232
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3232..3255
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             3262..3282
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3307..7407
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             3307..7407
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             7408..7455
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             7471..7527
                     /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             7528..8127
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(7528..7547)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     8024..8044
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    8143..8731
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8196..8216)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8614..8625)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             8803..9036
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    9114..9235
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(9151..9170)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     9205..9224
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      9275..9410
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(9430..9449)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(9431..9449)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(9459..9476)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(9459..9475)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9468..9490)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      9617..10072
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9704..9723)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9914..9935
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        10098..10202
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(10203..10271,10272..11063)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(10421..10440)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(11234..11590,1..232)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
       61 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
      121 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
      181 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
      241 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
      301 tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc
      361 tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc
      421 aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag
      481 gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca
      541 ttaggcaccc caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag
      601 cggataacaa tttcacacag gaaacagcta tgaccatgat tacgccaagc gcgcaattaa
      661 ccctcactaa agggaacaaa agctggagct gcaagcttaa tgtagtctta tgcaatactc
      721 ttgtagtctt gcaacatggt aacgatgagt tagcaacatg ccttacaagg agagaaaaag
      781 caccgtgcat gccgattggt ggaagtaagg tggtacgatc gtgccttatt aggaaggcaa
      841 cagacgggtc tgacatggat tggacgaacc actgaattgc cgcattgcag agatattgta
      901 tttaagtgcc tagctcgata cataaacggg tctctctggt tagaccagat ctgagcctgg
      961 gagctctctg gctaactagg gaacccactg cttaagcctc aataaagctt gccttgagtg
     1021 cttcaagtag tgtgtgcccg tctgttgtgt gactctggta actagagatc cctcagaccc
     1081 ttttagtcag tgtggaaaat ctctagcagt ggcgcccgaa cagggacttg aaagcgaaag
     1141 ggaaaccaga ggagctctct cgacgcagga ctcggcttgc tgaagcgcgc acggcaagag
     1201 gcgaggggcg gcgactggtg agtacgccaa aaattttgac tagcggaggc tagaaggaga
     1261 gagatgggtg cgagagcgtc agtattaagc gggggagaat tagatcgcga tgggaaaaaa
     1321 ttcggttaag gccaggggga aagaaaaaat ataaattaaa acatatagta tgggcaagca
     1381 gggagctaga acgattcgca gttaatcctg gcctgttaga aacatcagaa ggctgtagac
     1441 aaatactggg acagctacaa ccatcccttc agacaggatc agaagaactt agatcattat
     1501 ataatacagt agcaaccctc tattgtgtgc atcaaaggat agagataaaa gacaccaagg
     1561 aagctttaga caagatagag gaagagcaaa acaaaagtaa gaccaccgca cagcaagcgg
     1621 ccgctgatct tcagacctgg aggaggagat atgagggaca attggagaag tgaattatat
     1681 aaatataaag tagtaaaaat tgaaccatta ggagtagcac ccaccaaggc aaagagaaga
     1741 gtggtgcaga gagaaaaaag agcagtggga ataggagctt tgttccttgg gttcttggga
     1801 gcagcaggaa gcactatggg cgcagcgtca atgacgctga cggtacaggc cagacaatta
     1861 ttgtctggta tagtgcagca gcagaacaat ttgctgaggg ctattgaggc gcaacagcat
     1921 ctgttgcaac tcacagtctg gggcatcaag cagctccagg caagaatcct ggctgtggaa
     1981 agatacctaa aggatcaaca gctcctgggg atttggggtt gctctggaaa actcatttgc
     2041 accactgctg tgccttggaa tgctagttgg agtaataaat ctctggaaca gatttggaat
     2101 cacacgacct ggatggagtg ggacagagaa attaacaatt acacaagctt aatacactcc
     2161 ttaattgaag aatcgcaaaa ccagcaagaa aagaatgaac aagaattatt ggaattagat
     2221 aaatgggcaa gtttgtggaa ttggtttaac ataacaaatt ggctgtggta tataaaatta
     2281 ttcataatga tagtaggagg cttggtaggt ttaagaatag tttttgctgt actttctata
     2341 gtgaatagag ttaggcaggg atattcacca ttatcgtttc agacccacct cccaaccccg
     2401 aggggaccca gagagggcct atttcccatg attccttcat atttgcatat acgatacaag
     2461 gctgttagag agataattag aattaatttg actgtaaaca caaagatatt agtacaaaat
     2521 acgtgacgta gaaagtaata atttcttggg tagtttgcag ttttaaaatt atgttttaaa
     2581 atggactatc atatgcttac cgtaacttga aagtatttcg atttcttggc tttatatatc
     2641 ttgtggaaag gacgaaacac cgaccaccgt cagcactcag cggttttaga gctagaaata
     2701 gcaagttaaa ataaggctag tccgttatca acttgaaaaa gtggcaccga gtcggtgctt
     2761 ttttgaattc tagatcttga gacaaatggc agtattcatc cacaatttta aaagaaaagg
     2821 ggggattggg gggtacagtg caggggaaag aatagtagac ataatagcaa cagacataca
     2881 aactaaagaa ttacaaaaac aaattacaaa aattcaaaat tttcgggttt attacaggga
     2941 cagcagagat ccactttggc gccggctcga gtggctccgg tgcccgtcag tgggcagagc
     3001 gcacatcgcc cacagtcccc gagaagttgg ggggaggggt cggcaattga accggtgcct
     3061 agagaaggtg gcgcggggta aactgggaaa gtgatgtcgt gtactggctc cgcctttttc
     3121 ccgagggtgg gggagaaccg tatataagtg cagtagtcgc cgtgaacgtt ctttttcgca
     3181 acgggtttgc cgccagaaca caggtgtcgt gacgcgggat ccgccaccat ggattacaaa
     3241 gacgatgacg ataagatggc cccaaagaag aagcggaagg tcggtatcca cggagtccca
     3301 gcagccgaca agaagtacag catcggcctg gacatcggca ccaactctgt gggctgggcc
     3361 gtgatcaccg acgagtacaa ggtgcccagc aagaaattca aggtgctggg caacaccgac
     3421 cggcacagca tcaagaagaa cctgatcgga gccctgctgt tcgacagcgg cgaaacagcc
     3481 gaggccaccc ggctgaagag aaccgccaga agaagataca ccagacggaa gaaccggatc
     3541 tgctatctgc aagagatctt cagcaacgag atggccaagg tggacgacag cttcttccac
     3601 agactggaag agtccttcct ggtggaagag gataagaagc acgagcggca ccccatcttc
     3661 ggcaacatcg tggacgaggt ggcctaccac gagaagtacc ccaccatcta ccacctgaga
     3721 aagaaactgg tggacagcac cgacaaggcc gacctgcggc tgatctatct ggccctggcc
     3781 cacatgatca agttccgggg ccacttcctg atcgagggcg acctgaaccc cgacaacagc
     3841 gacgtggaca agctgttcat ccagctggtg cagacctaca accagctgtt cgaggaaaac
     3901 cccatcaacg ccagcggcgt ggacgccaag gccatcctgt ctgccagact gagcaagagc
     3961 agacggctgg aaaatctgat cgcccagctg cccggcgaga agaagaatgg cctgttcgga
     4021 aacctgattg ccctgagcct gggcctgacc cccaacttca agagcaactt cgacctggcc
     4081 gaggatgcca aactgcagct gagcaaggac acctacgacg acgacctgga caacctgctg
     4141 gcccagatcg gcgaccagta cgccgacctg tttctggccg ccaagaacct gtccgacgcc
     4201 atcctgctga gcgacatcct gagagtgaac accgagatca ccaaggcccc cctgagcgcc
     4261 tctatgatca agagatacga cgagcaccac caggacctga ccctgctgaa agctctcgtg
     4321 cggcagcagc tgcctgagaa gtacaaagag attttcttcg accagagcaa gaacggctac
     4381 gccggctaca ttgacggcgg agccagccag gaagagttct acaagttcat caagcccatc
     4441 ctggaaaaga tggacggcac cgaggaactg ctcgtgaagc tgaacagaga ggacctgctg
     4501 cggaagcagc ggaccttcga caacggcagc atcccccacc agatccacct gggagagctg
     4561 cacgccattc tgcggcggca ggaagatttt tacccattcc tgaaggacaa ccgggaaaag
     4621 atcgagaaga tcctgacctt ccgcatcccc tactacgtgg gccctctggc caggggaaac
     4681 agcagattcg cctggatgac cagaaagagc gaggaaacca tcaccccctg gaacttcgag
     4741 gaagtggtgg acaagggcgc ttccgcccag agcttcatcg agcggatgac caacttcgat
     4801 aagaacctgc ccaacgagaa ggtgctgccc aagcacagcc tgctgtacga gtacttcacc
     4861 gtgtataacg agctgaccaa agtgaaatac gtgaccgagg gaatgagaaa gcccgccttc
     4921 ctgagcggcg agcagaaaaa ggccatcgtg gacctgctgt tcaagaccaa ccggaaagtg
     4981 accgtgaagc agctgaaaga ggactacttc aagaaaatcg agtgcttcga ctccgtggaa
     5041 atctccggcg tggaagatcg gttcaacgcc tccctgggca cataccacga tctgctgaaa
     5101 attatcaagg acaaggactt cctggacaat gaggaaaacg aggacattct ggaagatatc
     5161 gtgctgaccc tgacactgtt tgaggacaga gagatgatcg aggaacggct gaaaacctat
     5221 gcccacctgt tcgacgacaa agtgatgaag cagctgaagc ggcggagata caccggctgg
     5281 ggcaggctga gccggaagct gatcaacggc atccgggaca agcagtccgg caagacaatc
     5341 ctggatttcc tgaagtccga cggcttcgcc aacagaaact tcatgcagct gatccacgac
     5401 gacagcctga cctttaaaga ggacatccag aaagcccagg tgtccggcca gggcgatagc
     5461 ctgcacgagc acattgccaa tctggccggc agccccgcca ttaagaaggg catcctgcag
     5521 acagtgaagg tggtggacga gctcgtgaaa gtgatgggcc ggcacaagcc cgagaacatc
     5581 gtgatcgaaa tggccagaga gaaccagacc acccagaagg gacagaagaa cagccgcgag
     5641 agaatgaagc ggatcgaaga gggcatcaaa gagctgggca gccagatcct gaaagaacac
     5701 cccgtggaaa acacccagct gcagaacgag aagctgtacc tgtactacct gcagaatggg
     5761 cgggatatgt acgtggacca ggaactggac atcaaccggc tgtccgacta cgatgtggac
     5821 catatcgtgc ctcagagctt tctgaaggac gactccatcg acaacaaggt gctgaccaga
     5881 agcgacaaga accggggcaa gagcgacaac gtgccctccg aagaggtcgt gaagaagatg
     5941 aagaactact ggcggcagct gctgaacgcc aagctgatta cccagagaaa gttcgacaat
     6001 ctgaccaagg ccgagagagg cggcctgagc gaactggata aggccggctt catcaagaga
     6061 cagctggtgg aaacccggca gatcacaaag cacgtggcac agatcctgga ctcccggatg
     6121 aacactaagt acgacgagaa tgacaagctg atccgggaag tgaaagtgat caccctgaag
     6181 tccaagctgg tgtccgattt ccggaaggat ttccagtttt acaaagtgcg cgagatcaac
     6241 aactaccacc acgcccacga cgcctacctg aacgccgtcg tgggaaccgc cctgatcaaa
     6301 aagtacccta agctggaaag cgagttcgtg tacggcgact acaaggtgta cgacgtgcgg
     6361 aagatgatcg ccaagagcga gcaggaaatc ggcaaggcta ccgccaagta cttcttctac
     6421 agcaacatca tgaacttttt caagaccgag attaccctgg ccaacggcga gatccggaag
     6481 cggcctctga tcgagacaaa cggcgaaacc ggggagatcg tgtgggataa gggccgggat
     6541 tttgccaccg tgcggaaagt gctgagcatg ccccaagtga atatcgtgaa aaagaccgag
     6601 gtgcagacag gcggcttcag caaagagtct atcctgccca agaggaacag cgataagctg
     6661 atcgccagaa agaaggactg ggaccctaag aagtacggcg gcttcgacag ccccaccgtg
     6721 gcctattctg tgctggtggt ggccaaagtg gaaaagggca agtccaagaa actgaagagt
     6781 gtgaaagagc tgctggggat caccatcatg gaaagaagca gcttcgagaa gaatcccatc
     6841 gactttctgg aagccaaggg ctacaaagaa gtgaaaaagg acctgatcat caagctgcct
     6901 aagtactccc tgttcgagct ggaaaacggc cggaagagaa tgctggcctc tgccggcgaa
     6961 ctgcagaagg gaaacgaact ggccctgccc tccaaatatg tgaacttcct gtacctggcc
     7021 agccactatg agaagctgaa gggctccccc gaggataatg agcagaaaca gctgtttgtg
     7081 gaacagcaca agcactacct ggacgagatc atcgagcaga tcagcgagtt ctccaagaga
     7141 gtgatcctgg ccgacgctaa tctggacaaa gtgctgtccg cctacaacaa gcaccgggat
     7201 aagcccatca gagagcaggc cgagaatatc atccacctgt ttaccctgac caatctggga
     7261 gcccctgccg ccttcaagta ctttgacacc accatcgacc ggaagaggta caccagcacc
     7321 aaagaggtgc tggacgccac cctgatccac cagagcatca ccggcctgta cgagacacgg
     7381 atcgacctgt ctcagctggg aggcgacaag cgtcctgctg ctactaagaa agctggtcaa
     7441 gctaagaaaa agaaagctag cggcagcggc gccaccaact tcagcctgct gaagcaggcc
     7501 ggcgacgtgg aggagaaccc cggccccatg accgagtaca agcccacggt gcgcctcgcc
     7561 acccgcgacg acgtccccag ggccgtacgc accctcgccg ccgcgttcgc cgactacccc
     7621 gccacgcgcc acaccgtcga tccggaccgc cacatcgagc gggtcaccga gctgcaagaa
     7681 ctcttcctca cgcgcgtcgg gctcgacatc ggcaaggtgt gggtcgcgga cgacggcgcc
     7741 gcggtggcgg tctggaccac gccggagagc gtcgaagcgg gggcggtgtt cgccgagatc
     7801 ggcccgcgca tggccgagtt gagcggttcc cggctggccg cgcagcaaca gatggaaggc
     7861 ctcctggcgc cgcaccggcc caaggagccc gcgtggttcc tggccaccgt cggagtctcg
     7921 cccgaccacc agggcaaggg tctgggcagc gccgtcgtgc tccccggagt ggaggcggcc
     7981 gagcgcgccg gggtgcccgc cttcctggag acctccgcgc cccgcaacct ccccttctac
     8041 gagcggctcg gcttcaccgt caccgccgac gtcgaggtgc ccgaaggacc gcgcacctgg
     8101 tgcatgaccc gcaagcccgg tgcctgaacg cgttaagtcg acaatcaacc tctggattac
     8161 aaaatttgtg aaagattgac tggtattctt aactatgttg ctccttttac gctatgtgga
     8221 tacgctgctt taatgccttt gtatcatgct attgcttccc gtatggcttt cattttctcc
     8281 tccttgtata aatcctggtt gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa
     8341 cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca ctggttgggg cattgccacc
     8401 acctgtcagc tcctttccgg gactttcgct ttccccctcc ctattgccac ggcggaactc
     8461 atcgccgcct gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc
     8521 gtggtgttgt cggggaaatc atcgtccttt ccttggctgc tcgcctgtgt tgccacctgg
     8581 attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc ggaccttcct
     8641 tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg
     8701 agtcggatct ccctttgggc cgcctccccg cgtcgacttt aagaccaatg acttacaagg
     8761 cagctgtaga tcttagccac tttttaaaag aaaagggggg actggaaggg ctaattcact
     8821 cccaacgaag acaagatctg ctttttgctt gtactgggtc tctctggtta gaccagatct
     8881 gagcctggga gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc
     8941 cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc
     9001 tcagaccctt ttagtcagtg tggaaaatct ctagcagtac gtatagtagt tcatgtcatc
     9061 ttattattca gtatttataa cttgcaaaga aatgaatatc agagagtgag aggaacttgt
     9121 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     9181 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg
     9241 tctggctcta gctatcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
     9301 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     9361 ctcggcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc tagggacgta
     9421 cccaattcgc cctatagtga gtcgtattac gcgcgctcac tggccgtcgt tttacaacgt
     9481 cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc
     9541 gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc
     9601 ctgaatggcg aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     9661 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
     9721 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     9781 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat
     9841 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     9901 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc
     9961 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
    10021 atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat ttaggtggca
    10081 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
    10141 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
    10201 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
    10261 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
    10321 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
    10381 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
    10441 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
    10501 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
    10561 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
    10621 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
    10681 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
    10741 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
    10801 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
    10861 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
    10921 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
    10981 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
    11041 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
    11101 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
    11161 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
    11221 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
    11281 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
    11341 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
    11401 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
    11461 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
    11521 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
    11581 tggagcgaac
//
