LOCUS       sequence-466841-       12820 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  knockout Pnpt1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12820)
  AUTHORS   Miao R, Jiang C, Chang WY, Zhang H, An J, Ho F, Chen P, Zhang H, 
            Junqueira C, Amgalan D, Liang FG, Zhang J, Evavold CL, 
            Hafner-Bratkovic I, Zhang Z, Fontana P, Xia S, Waldeck-Weiermair M, 
            Pan Y, Michel T, Bar-Peled L, Wu H, Kagan JC, Kitsis RN, Zhang P, 
            Liu X, Lieberman J
  TITLE     Gasdermin D permeabilization of mitochondrial inner and outer 
            membranes accelerates and enhances pyroptosis.
  JOURNAL   Immunity. 2023 Nov 14;56(11):2523-2541.e8. doi: 
            10.1016/j.immuni.2023.10.004. Epub 2023 Nov 3.
  PUBMED    37924812
REFERENCE   2  (bases 1 to 12820)
  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"; doi: 
            "10.1016/j.immuni.2023.10.004"; journalName: "Immunity"; date: 
            "2023-11-14- 14"; volume: "56"; issue: "11"; pages: "2523-2541.e8"
COMMENT     Sequence Label: lentiCRISPR v2-sgPnpt1-1#
FEATURES             Location/Qualifiers
     source          1..12820
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(67..111)
                     /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    296..529
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     misc_feature    1022..1147
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             1194..1374
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     promoter        complement(1392..1590)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(1417..1437)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        1592..1971
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     2143..2162
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        2237..2341
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2342..2410,2411..3202)
                     /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(2560..2579)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3373..3961
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3862..3881
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4115..4132
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4249..4270
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4285..4290,4291..4308,4309..4315)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4323..4339
                     /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     4328..4350
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4347..4363
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4347..4363
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    4412..4533
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4423..4442)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4477..4496
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(4663..5037)
                     /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"
     promoter        complement(5056..5103)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(5151..5480)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5165..5300
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5219..5238)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     5465..5485
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(5494..5922)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5604..5625)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5816..5835
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    5968..6192
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     6181..6198
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     LTR             6224..6457
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     misc_feature    6529..7117
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             6635..6646
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     7044..7064
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(7142..7540)
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     CDS             complement(7541..7597)
                     /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             complement(7607..7630)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             complement(7631..7678)
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             complement(7679..11782)
                     /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             complement(7679..11782)
                     /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"
     regulatory      11779..11788
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(11807..12018)
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     misc_RNA        complement(12080..12155)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(12173..12421)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(12231..12250)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(12401..12421)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     misc_feature    12472..12589
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 gcctcctact atcattatga ataattttat ataccacagc caatttgtta tgttaaacca
       61 attccacaaa cttgcccatt tatctaattc caataattct tgttcattct tttcttgctg
      121 gttttgcgat tcttcaatta aggagtgtat taagcttgtg taattgttaa tttctctgtc
      181 ccactccatc caggtcgtgt gattccaaat ctgttccaga gatttattac tccaactagc
      241 attccaaggc acagcagtgg tgcaaatgag ttttccagag caaccccaaa tccccaggag
      301 ctgttgatcc tttaggtatc tttccacagc caggattctt gcctggagct gcttgatgcc
      361 ccagactgtg agttgcaaca gatgctgttg cgcctcaata gccctcagca aattgttctg
      421 ctgctgcact ataccagaca ataattgtct ggcctgtacc gtcagcgtca ttgacgctgc
      481 gcccatagtg cttcctgctg ctcccaagaa cccaaggaac aaagctccta ttcccactgc
      541 tcttttttct ctctgcacca ctcttctctt tgccttggtg ggtgctactc ctaatggttc
      601 aatttttact actttatatt tatataattc acttctccaa ttgtccctca tatctcctcc
      661 tccaggtctg aagatcagcg gccgcttgct gtgcggtggt cttacttttg ttttgctctt
      721 cctctatctt gtctaaagct tccttggtgt cttttatctc tatcctttga tgcacacaat
      781 agagggttgc tactgtatta tataatgatc taagttcttc tgatcctgtc tgaagggatg
      841 gttgtagctg tcccagtatt tgtctacagc cttctgatgt ttctaacagg ccaggattaa
      901 ctgcgaatcg ttctagctcc ctgcttgccc atactatatg ttttaattta tattttttct
      961 ttccccctgg ccttaaccga attttttccc atcgcgatct aattctcccc cgcttaatac
     1021 tgacgctctc gcacccatct ctctccttct agcctccgct agtcaaaatt tttggcgtac
     1081 tcaccagtcg ccgcccctcg cctcttgccg tgcgcgcttc agcaagccga gtcctgcgtc
     1141 gagagagctc ctctggtttc cctttcgctt tcaagtccct gttcgggcgc cactgctaga
     1201 gattttccac actgactaaa agggtctgag ggatctctag ttaccagagt cacacaacag
     1261 acgggcacac actacttgaa gcactcaagg caagctttat tgaggcttaa gcagtgggtt
     1321 ccctagttag ccagagagct cccaggctca gatctggtct aaccagagag acccagtaca
     1381 ggcaaaacgc gctgcttata tagacctccc accgtacacg cctaccgccc atttgcgtca
     1441 atggggcgga gttgttacga cattttggaa agtcccgttg attttggtgc caaaacaaac
     1501 tcccattgac gtcaatgggg tggagacttg gaaatccccg tgagtcaaac cgctatccac
     1561 gcccattgat gtactgccaa aaccgcatca ccatggtaat agcgatgact aatacgtaga
     1621 tgtactgcca agtaggaaag tcccataagg tcatgtactg ggcataatgc caggcgggcc
     1681 atttaccgtc attgacgtca atagggggcg tacttggcat atgatacact tgatgtactg
     1741 ccaagtgggc agtttaccgt aaatactcca cccattgacg tcaatggaaa gtccctattg
     1801 gcgttactat gggaacatac gtcattattg acgtcaatgg gcgggggtcg ttgggcggtc
     1861 agccaggcgg gccatttacc gtaagttatg taacgcggaa ctccatatat gggctatgaa
     1921 ctaatgaccc cgtaattgat tactattaat aactagtcaa taatcaatgt caacgcgtat
     1981 atctggcccg tacatcgcga agcagcgcaa aacgcctaac cctaagcaga ttcttcatgc
     2041 aattgtcggt caagccttgc cttgttgtag cttaaatttt gctcgcgcac tactcagcga
     2101 cctccaacac acaagcaggg agcagatact ggcttaacta tgcggcatca gagcagattg
     2161 tactgagagt gcaccatagg ggatcgggag atctcccgat ccgtcgacgt caggtggcac
     2221 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     2281 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     2341 tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc
     2401 tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc
     2461 acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc
     2521 cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc
     2581 ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt
     2641 ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt
     2701 atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat
     2761 cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct
     2821 tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat
     2881 gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc
     2941 ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg
     3001 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc
     3061 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
     3121 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
     3181 ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga
     3241 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     3301 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     3361 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     3421 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     3481 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt
     3541 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     3601 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     3661 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     3721 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     3781 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     3841 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     3901 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     3961 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     4021 gttctttcct gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc
     4081 tgataccgct cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga
     4141 agagcgccca atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg
     4201 gcacgacagg tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta
     4261 gctcactcat taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg
     4321 aattgtgagc ggataacaat ttcacacagg aaacagctat gaccatgatt acgccaagct
     4381 ctagctagag gtcgacggta tacagacatg ataagataca ttgatgagtt tggacaaacc
     4441 acaactagaa tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta
     4501 tttgtaacca ttataagctg caataaacaa gttggggtgg gcgaagaact ccagcatgag
     4561 atccccgcgc tggaggatca tccagccggc gtcccggaaa acgattccga agcccaacct
     4621 ttcatagaag gcggcggtgg aatcgaaatc tcgtagcacg tgtcagtcct gctcctcggc
     4681 cacgaagtgc acgcagttgc cggccgggtc gcgcagggcg aactcccgcc cccacggctg
     4741 ctcgccgatc tcggtcatgg ccggcccgga ggcgtcccgg aagttcgtgg acacgacctc
     4801 cgaccactcg gcgtacagct cgtccaggcc gcgcacccac acccaggcca gggtgttgtc
     4861 cggcaccacc tggtcctgga ccgcgctgat gaacagggtc acgtcgtccc ggaccacacc
     4921 ggcgaagtcg tcctccacga agtcccggga gaacccgagc cggtcggtcc agaactcgac
     4981 cgctccggcg acgtcgcgcg cggtgagcac cggaacggca ctggtcaact tggccatggt
     5041 ttagttcctc accttgtcgt attatactat gccgatatac tatgccgatg attaattgtc
     5101 aacacgtgct gatcagatcc gaaaatggat atacaagctc ccgggagctt tttgcaaaag
     5161 cctaggcctc caaaaaagcc tcctcactac ttctggaata gctcagaggc agaggcggcc
     5221 tcggcctctg cataaataaa aaaaattagt cagccatggg gcggagaatg ggcggaactg
     5281 ggcggagtta ggggcgggat gggcggagtt aggggcggga ctatggttgc tgactaattg
     5341 agatgcatgc tttgcatact tctgcctgct ggggagcctg gggactttcc acacctggtt
     5401 gctgactaat tgagatgcat gctttgcata cttctgcctg ctggggagcc tggggacttt
     5461 ccacacccta actgacacac attccacaga attaattcgc gttaaatttt tgttaaatca
     5521 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
     5581 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
     5641 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
     5701 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
     5761 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga
     5821 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
     5881 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtggggatac cccctagagc
     5941 cccagctggt tctttccgcc tcagaagcca tagagcccac cgcatcccca gcatgcctgc
     6001 tattgtcttc ccaatcctcc cccttgctgt cctgccccac cccacccccc agaatagaat
     6061 gacacctact cagacaatgc gatgcaattt cctcatttta ttaggaaagg acagtgggag
     6121 tggcaccttc cagggtcaag gaaggcacgg gggaggggca aacaacagat ggctggcaac
     6181 tagaaggcac agtcgaggct gatcagcggg tttaaacggg ccctgctaga gattttccac
     6241 actgactaaa agggtctgag ggatctctag ttaccagagt cacacaacag acgggcacac
     6301 actacttgaa gcactcaagg caagctttat tgaggcttaa gcagtgggtt ccctagttag
     6361 ccagagagct cccaggctca gatctggtct aaccagagag acccagtaca agcaaaaagc
     6421 agatcttgtc ttcgttggga gtgaattagc ccttccagtc cccccttttc ttttaaaaag
     6481 tggctaagat ctacagctgc cttgtaagtc attggtctta aagtcgacgc ggggaggcgg
     6541 cccaaaggga gatccgactc gtctgagggc gaaggcgaag acgcggaaga ggccgcagag
     6601 ccggcagcag gccgcgggaa ggaaggtccg ctggattgag ggccgaaggg acgtagcaga
     6661 aggacgtccc gcgcagaatc caggtggcaa cacaggcgag cagccaagga aaggacgatg
     6721 atttccccga caacaccacg gaattgtcag tgcccaacag ccgagcccct gtccagcagc
     6781 gggcaaggca ggcggcgatg agttccgccg tggcaatagg gagggggaaa gcgaaagtcc
     6841 cggaaaggag ctgacaggtg gtggcaatgc cccaaccagt gggggttgcg tcagcaaaca
     6901 cagtgcacac cacgccacgt tgcctgacaa cgggccacaa ctcctcataa agagacagca
     6961 accaggattt atacaaggag gagaaaatga aagccatacg ggaagcaata gcatgataca
     7021 aaggcattaa agcagcgtat ccacatagcg taaaaggagc aacatagtta agaataccag
     7081 tcaatctttc acaaattttg taatccagag gttgattacc gataagcttg atatcgaatt
     7141 cttagccctc ccacacataa ccagagggca gcaattcacg aatcccaact gccgtcggct
     7201 gtccatcact gtccttcact atggctttga tcccaggatg cagatcgaga agcacctgtc
     7261 ggcaccgtcc gcaggggctc aagatgcccc tgttctcatt tccgatcgcg acgatacaag
     7321 tcaggttgcc agctgccgca gcagcagcag tgcccagcac cacgagttct gcacaaggtc
     7381 ccccagtaaa atgatataca ttgacaccag tgaagatgcg gccgtcgcta gagagagctg
     7441 cgctggcgac gctgtagtct tcagagatgg ggatgctgtt gattgtagcc gttgctcttt
     7501 caatgagggt ggattcttct tgagacaaag gcttggccat cggtccagga ttctcttcga
     7561 catctccggc ttgtttcagc agagagaagt ttgttgcgcc ggatccctta tcgtcatcgt
     7621 ctttgtaatc tttcttcttc ttagcctgtc cagccttctt tgtggcggca ggtcgcttgt
     7681 cgcctcccag ctgagacagg tcgatccgtg tctcgtacag gccggtgatg ctctggtgga
     7741 tcagggtggc gtccagcacc tctttggtgc tggtgtacct cttccggtcg atggtggtgt
     7801 caaagtactt gaaggcggca ggggctccca gattggtcag ggtaaacagg tggatgatat
     7861 tctcggcctg ctctctgatg ggcttatccc ggtgcttgtt gtaggcggac agcactttgt
     7921 ccagattagc gtcggccagg atcactctct tggagaactc gctgatctgc tcgatgatct
     7981 cgtccaggta gtgcttgtgc tgttccacaa acagctgttt ctgctcatta tcctcggggg
     8041 agcccttcag cttctcatag tggctggcca ggtacaggaa gttcacatat ttggagggca
     8101 gggccagttc gtttcccttc tgcagttcgc cggcagaggc cagcattctc ttccggccgt
     8161 tttccagctc gaacagggag tacttaggca gcttgatgat caggtccttt ttcacttctt
     8221 tgtagccctt ggcttccaga aagtcgatgg gattcttctc gaagctgctt ctttccatga
     8281 tggtgatccc cagcagctct ttcacactct tcagtttctt ggacttgccc ttttccactt
     8341 tggccaccac cagcacagaa taggccacgg tggggctgtc gaagccgccg tacttcttag
     8401 ggtcccagtc cttctttctg gcgatcagct tatcgctgtt cctcttgggc aggatagact
     8461 ctttgctgaa gccgcctgtc tgcacctcgg tctttttcac gatattcact tggggcatgc
     8521 tcagcacttt ccgcacggtg gcaaaatccc ggcccttatc ccacacgatc tccccggttt
     8581 cgccgtttgt ctcgatcaga ggccgcttcc ggatctcgcc gttggccagg gtaatctcgg
     8641 tcttgaaaaa gttcatgatg ttgctgtaga agaagtactt ggcggtagcc ttgccgattt
     8701 cctgctcgct cttggcgatc atcttccgca cgtcgtacac cttgtagtcg ccgtacacga
     8761 actcgctttc cagcttaggg tactttttga tcagggcggt tcccacgacg gcgttcaggt
     8821 aggcgtcgtg ggcgtggtgg tagttgttga tctcgcgcac tttgtaaaac tggaaatcct
     8881 tccggaaatc ggacaccagc ttggacttca gggtgatcac tttcacttcc cggatcagct
     8941 tgtcattctc gtcgtactta gtgttcatcc gggagtccag gatctgtgcc acgtgctttg
     9001 tgatctgccg ggtttccacc agctgtctct tgatgaagcc ggccttatcc agttcgctca
     9061 ggccgcctct ctcggccttg gtcagattgt cgaactttct ctgggtaatc agcttggcgt
     9121 tcagcagctg ccgccagtag ttcttcatct tcttcacgac ctcttcggag ggcacgttgt
     9181 cgctcttgcc ccggttcttg tcgcttctgg tcagcacctt gttgtcgatg gagtcgtcct
     9241 tcagaaagct ctgaggcacg atatggtcca catcgtagtc ggacagccgg ttgatgtcca
     9301 gttcctggtc cacgtacata tcccgcccat tctgcaggta gtacaggtac agcttctcgt
     9361 tctgcagctg ggtgttttcc acggggtgtt ctttcaggat ctggctgccc agctctttga
     9421 tgccctcttc gatccgcttc attctctcgc ggctgttctt ctgtcccttc tgggtggtct
     9481 ggttctctct ggccatttcg atcacgatgt tctcgggctt gtgccggccc atcactttca
     9541 cgagctcgtc caccaccttc actgtctgca ggatgccctt cttaatggcg gggctgccgg
     9601 ccagattggc aatgtgctcg tgcaggctat cgccctggcc ggacacctgg gctttctgga
     9661 tgtcctcttt aaaggtcagg ctgtcgtcgt ggatcagctg catgaagttt ctgttggcga
     9721 agccgtcgga cttcaggaaa tccaggattg tcttgccgga ctgcttgtcc cggatgccgt
     9781 tgatcagctt ccggctcagc ctgccccagc cggtgtatct ccgccgcttc agctgcttca
     9841 tcactttgtc gtcgaacagg tgggcatagg ttttcagccg ttcctcgatc atctctctgt
     9901 cctcaaacag tgtcagggtc agcacgatat cttccagaat gtcctcgttt tcctcattgt
     9961 ccaggaagtc cttgtccttg ataattttca gcagatcgtg gtatgtgccc agggaggcgt
    10021 tgaaccgatc ttccacgccg gagatttcca cggagtcgaa gcactcgatt ttcttgaagt
    10081 agtcctcttt cagctgcttc acggtcactt tccggttggt cttgaacagc aggtccacga
    10141 tggccttttt ctgctcgccg ctcaggaagg cgggctttct cattccctcg gtcacgtatt
    10201 tcactttggt cagctcgtta tacacggtga agtactcgta cagcaggctg tgcttgggca
    10261 gcaccttctc gttgggcagg ttcttatcga agttggtcat ccgctcgatg aagctctggg
    10321 cggaagcgcc cttgtccacc acttcctcga agttccaggg ggtgatggtt tcctcgctct
    10381 ttctggtcat ccaggcgaat ctgctgtttc ccctggccag agggcccacg tagtagggga
    10441 tgcggaaggt caggatcttc tcgatctttt cccggttgtc cttcaggaat gggtaaaaat
    10501 cttcctgccg ccgcagaatg gcgtgcagct ctcccaggtg gatctggtgg gggatgctgc
    10561 cgttgtcgaa ggtccgctgc ttccgcagca ggtcctctct gttcagcttc acgagcagtt
    10621 cctcggtgcc gtccatcttt tccaggatgg gcttgatgaa cttgtagaac tcttcctggc
    10681 tggctccgcc gtcaatgtag ccggcgtagc cgttcttgct ctggtcgaag aaaatctctt
    10741 tgtacttctc aggcagctgc tgccgcacga gagctttcag cagggtcagg tcctggtggt
    10801 gctcgtcgta tctcttgatc atagaggcgc tcaggggggc cttggtgatc tcggtgttca
    10861 ctctcaggat gtcgctcagc aggatggcgt cggacaggtt cttggcggcc agaaacaggt
    10921 cggcgtactg gtcgccgatc tgggccagca ggttgtccag gtcgtcgtcg taggtgtcct
    10981 tgctcagctg cagtttggca tcctcggcca ggtcgaagtt gctcttgaag ttgggggtca
    11041 ggcccaggct cagggcaatc aggtttccga acaggccatt cttcttctcg ccgggcagct
    11101 gggcgatcag attttccagc cgtctgctct tgctcagtct ggcagacagg atggccttgg
    11161 cgtccacgcc gctggcgttg atggggtttt cctcgaacag ctggttgtag gtctgcacca
    11221 gctggatgaa cagcttgtcc acgtcgctgt tgtcggggtt caggtcgccc tcgatcagga
    11281 agtggccccg gaacttgatc atgtgggcca gggccagata gatcagccgc aggtcggcct
    11341 tgtcggtgct gtccaccagt ttctttctca ggtggtagat ggtggggtac ttctcgtggt
    11401 aggccacctc gtccacgatg ttgccgaaga tggggtgccg ctcgtgcttc ttatcctctt
    11461 ccaccaggaa ggactcttcc agtctgtgga agaagctgtc gtccaccttg gccatctcgt
    11521 tgctgaagat ctcttgcaga tagcagatcc ggttcttccg tctggtgtat cttcttctgg
    11581 cggttctctt cagccgggtg gcctcggctg tttcgccgct gtcgaacagc agggctccga
    11641 tcaggttctt cttgatgctg tgccggtcgg tgttgcccag caccttgaat ttcttgctgg
    11701 gcaccttgta ctcgtcggtg atcacggccc agcccacaga gttggtgccg atgtccaggc
    11761 cgatgctgta cttcttgtcc atggtggcag cgctctagaa ccggtcctgt gttctggcgg
    11821 caaacccgtt gcgaaaaaga acgttcacgg cgactactgc acttatatac ggttctcccc
    11881 caccctcggg aaaaaggcgg agccagtaca cgacatcact ttcccagttt accccgcgcc
    11941 accttctcta ggcaccggat caattgccga cccctccccc caacttctcg gggactgtgg
    12001 gcgatgtgcg ctctgcccac tgacgggcac cggagccaat tcccactcct ttcaagacct
    12061 agctagcgaa ttcaaaaaag caccgactcg gtgccacttt ttcaagttga taacggacta
    12121 gccttatttt aacttgctat ttctagctct aaaactgttg ggttaacgac acggtgtttc
    12181 gtcctttcca caagatatat aaagccaaga aatcgaaata ctttcaagtt acggtaagca
    12241 tatgatagtc cattttaaaa cataatttta aaactgcaaa ctacccaaga aattattact
    12301 ttctacgtca cgtattttgt actaatatct ttgtgtttac agtcaaatta attctaatta
    12361 tctctctaac agccttgtat cgtatatgca aatatgaagg aatcatggga aataggccct
    12421 cggtacctta attaaccaaa ctggatctct gctgtccctg taataaaccc gaaaattttg
    12481 aatttttgta atttgttttt gtaattcttt agtttgtatg tctgttgcta ttatgtctac
    12541 tattctttcc cctgcactgt accccccaat cccccctttt cttttaaaat tgtggatgaa
    12601 tactgccatt tgtctgcaga attggcgcac gcagtgccga tccgttcact aatcgaatgg
    12661 atctgtctct gtctctctct ccaccttctt cttctattcc ttcgggcctg tcgggtcccc
    12721 tcggggttgg gaggtgggtc tgaaacgata atggtgaata tccctgccta actctattca
    12781 ctatagaaag tacagcaaaa actattctta aacctaccaa
//
