LOCUS       sequence-460796-       13400 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Knockout mouse Il1r1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13400)
  AUTHORS   Hanggi K, Li J, Gangadharan A, Liu X, Celias DP, Osunmakinde O, 
            Keske A, Davis J, Ahmad F, Giron A, Anadon CM, Gardner A, DeNardo 
            DG, Shaw TI, Beg AA, Yu X, Ruffell B
  TITLE     Interleukin-1alpha release during necrotic-like cell death generates
            myeloid-driven immunosuppression that restricts anti-tumor immunity.
  JOURNAL   Cancer Cell. 2024 Nov 13:S1535-6108(24)00402-1. doi: 
            10.1016/j.ccell.2024.10.014.
  PUBMED    39577420
REFERENCE   2  (bases 1 to 13400)
  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: "Cancer 
            Cell. 2024 Nov 13:S1535-6108(24)00402-1. doi: 
            10.1016/j.ccell.2024.10.014."
COMMENT     Sequence Label: pLenti-CRISPRv2-sgIl1r1_#2-hygro
FEATURES             Location/Qualifiers
     source          1..13400
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(353..397)
                     /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    582..815
                     /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    1308..1433
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             1480..1660
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     promoter        complement(1678..1876)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(1703..1723)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        1878..2257
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     2433..2452
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        2527..2631
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2632..2700,2701..3492)
                     /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(2850..2869)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3663..4251
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4152..4171
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4405..4422
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4539..4560
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4575..4580,4581..4598,4599..4605)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4613..4629
                     /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     4618..4640
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4637..4653
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4637..4653
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    4702..4823
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4713..4732)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4767..4786
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(4953..5327)
                     /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(5346..5393)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(5441..5770)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5455..5590
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5509..5528)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     5755..5775
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(5784..6212)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5894..5915)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6106..6125
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    6258..6482
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     6471..6488
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     LTR             6514..6747
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     misc_feature    6819..7407
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             6925..6936
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     7334..7354
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8404..8460)
                     /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(8470..8493)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             complement(8494..8541)
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             complement(8542..12645)
                     /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(8542..12645)
                     /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      12642..12651
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(12670..12881)
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     misc_RNA        complement(12943..13018)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(13039..13287)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(13097..13116)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(13267..13287)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     misc_feature    join(13338..13400,1..55)
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 gtctactatt ctttcccctg cactgtaccc cccaatcccc ccttttcttt taaaattgtg
       61 gatgaatact gccatttgtc tgcagaattg gcgcacgcag tgccgatccg ttcactaatc
      121 gaatggatct gtctctgtct ctctctccac cttcttcttc tattccttcg ggcctgtcgg
      181 gtcccctcgg ggttgggagg tgggtctgaa acgataatgg tgaatatccc tgcctaactc
      241 tattcactat agaaagtaca gcaaaaacta ttcttaaacc taccaagcct cctactatca
      301 ttatgaataa ttttatatac cacagccaat ttgttatgtt aaaccaattc cacaaacttg
      361 cccatttatc taattccaat aattcttgtt cattcttttc ttgctggttt tgcgattctt
      421 caattaagga gtgtattaag cttgtgtaat tgttaatttc tctgtcccac tccatccagg
      481 tcgtgtgatt ccaaatctgt tccagagatt tattactcca actagcattc caaggcacag
      541 cagtggtgca aatgagtttt ccagagcaac cccaaatccc caggagctgt tgatccttta
      601 ggtatctttc cacagccagg attcttgcct ggagctgctt gatgccccag actgtgagtt
      661 gcaacagatg ctgttgcgcc tcaatagccc tcagcaaatt gttctgctgc tgcactatac
      721 cagacaataa ttgtctggcc tgtaccgtca gcgtcattga cgctgcgccc atagtgcttc
      781 ctgctgctcc caagaaccca aggaacaaag ctcctattcc cactgctctt ttttctctct
      841 gcaccactct tctctttgcc ttggtgggtg ctactcctaa tggttcaatt tttactactt
      901 tatatttata taattcactt ctccaattgt ccctcatatc tcctcctcca ggtctgaaga
      961 tcagcggccg cttgctgtgc ggtggtctta cttttgtttt gctcttcctc tatcttgtct
     1021 aaagcttcct tggtgtcttt tatctctatc ctttgatgca cacaatagag ggttgctact
     1081 gtattatata atgatctaag ttcttctgat cctgtctgaa gggatggttg tagctgtccc
     1141 agtatttgtc tacagccttc tgatgtttct aacaggccag gattaactgc gaatcgttct
     1201 agctccctgc ttgcccatac tatatgtttt aatttatatt ttttctttcc ccctggcctt
     1261 aaccgaattt tttcccatcg cgatctaatt ctcccccgct taatactgac gctctcgcac
     1321 ccatctctct ccttctagcc tccgctagtc aaaatttttg gcgtactcac cagtcgccgc
     1381 ccctcgcctc ttgccgtgcg cgcttcagca agccgagtcc tgcgtcgaga gagctcctct
     1441 ggtttccctt tcgctttcaa gtccctgttc gggcgccact gctagagatt ttccacactg
     1501 actaaaaggg tctgagggat ctctagttac cagagtcaca caacagacgg gcacacacta
     1561 cttgaagcac tcaaggcaag ctttattgag gcttaagcag tgggttccct agttagccag
     1621 agagctccca ggctcagatc tggtctaacc agagagaccc agtacaggca aaacgcgctg
     1681 cttatataga cctcccaccg tacacgccta ccgcccattt gcgtcaatgg ggcggagttg
     1741 ttacgacatt ttggaaagtc ccgttgattt tggtgccaaa acaaactccc attgacgtca
     1801 atggggtgga gacttggaaa tccccgtgag tcaaaccgct atccacgccc attgatgtac
     1861 tgccaaaacc gcatcaccat ggtaatagcg atgactaata cgtagatgta ctgccaagta
     1921 ggaaagtccc ataaggtcat gtactgggca taatgccagg cgggccattt accgtcattg
     1981 acgtcaatag ggggcgtact tggcatatga tacacttgat gtactgccaa gtgggcagtt
     2041 taccgtaaat actccaccca ttgacgtcaa tggaaagtcc ctattggcgt tactatggga
     2101 acatacgtca ttattgacgt caatgggcgg gggtcgttgg gcggtcagcc aggcgggcca
     2161 tttaccgtaa gttatgtaac gcggaactcc atatatgggc tatgaactaa tgaccccgta
     2221 attgattact attaataact agtcaataat caatgtcaac gcgcgcgtat atctggcccg
     2281 tacatcgcga agcagcgcaa aacgcctaac cctaagcaga ttcttcatgc aattgtcggt
     2341 caagccttgc cttgttgtag cttaaatttt gctcgcgcac tactcagcga cctccaacac
     2401 acaagcaggg agcagatact ggcttaacta tgcggcatca gagcagattg tactgagagt
     2461 gcaccatagg ggatcgggag atctcccgat ccgtcgacgt caggtggcac ttttcgggga
     2521 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     2581 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     2641 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     2701 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     2761 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     2821 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     2881 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     2941 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     3001 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     3061 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     3121 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     3181 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     3241 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     3301 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     3361 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     3421 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     3481 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     3541 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     3601 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     3661 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     3721 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     3781 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     3841 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     3901 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     3961 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     4021 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     4081 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     4141 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     4201 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     4261 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
     4321 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct
     4381 cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgccca
     4441 atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg
     4501 tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta gctcactcat
     4561 taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc
     4621 ggataacaat ttcacacagg aaacagctat gaccatgatt acgccaagct ctagctagag
     4681 gtcgacggta tacagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa
     4741 tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca
     4801 ttataagctg caataaacaa gttggggtgg gcgaagaact ccagcatgag atccccgcgc
     4861 tggaggatca tccagccggc gtcccggaaa acgattccga agcccaacct ttcatagaag
     4921 gcggcggtgg aatcgaaatc tcgtagcacg tgtcagtcct gctcctcggc cacgaagtgc
     4981 acgcagttgc cggccgggtc gcgcagggcg aactcccgcc cccacggctg ctcgccgatc
     5041 tcggtcatgg ccggcccgga ggcgtcccgg aagttcgtgg acacgacctc cgaccactcg
     5101 gcgtacagct cgtccaggcc gcgcacccac acccaggcca gggtgttgtc cggcaccacc
     5161 tggtcctgga ccgcgctgat gaacagggtc acgtcgtccc ggaccacacc ggcgaagtcg
     5221 tcctccacga agtcccggga gaacccgagc cggtcggtcc agaactcgac cgctccggcg
     5281 acgtcgcgcg cggtgagcac cggaacggca ctggtcaact tggccatggt ttagttcctc
     5341 accttgtcgt attatactat gccgatatac tatgccgatg attaattgtc aacacgtgct
     5401 gatcagatcc gaaaatggat atacaagctc ccgggagctt tttgcaaaag cctaggcctc
     5461 caaaaaagcc tcctcactac ttctggaata gctcagaggc agaggcggcc tcggcctctg
     5521 cataaataaa aaaaattagt cagccatggg gcggagaatg ggcggaactg ggcggagtta
     5581 ggggcgggat gggcggagtt aggggcggga ctatggttgc tgactaattg agatgcatgc
     5641 tttgcatact tctgcctgct ggggagcctg gggactttcc acacctggtt gctgactaat
     5701 tgagatgcat gctttgcata cttctgcctg ctggggagcc tggggacttt ccacacccta
     5761 actgacacac attccacaga attaattcgc gttaaatttt tgttaaatca gctcattttt
     5821 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
     5881 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt
     5941 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc
     6001 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
     6061 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
     6121 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
     6181 cgccgcgctt aatgcgccgc tacagggcgc gtggggatac cccctagagc cccagctggt
     6241 tctttccgcc tcagaagcca tagagcccac cgcatcccca gcatgcctgc tattgtcttc
     6301 ccaatcctcc cccttgctgt cctgccccac cccacccccc agaatagaat gacacctact
     6361 cagacaatgc gatgcaattt cctcatttta ttaggaaagg acagtgggag tggcaccttc
     6421 cagggtcaag gaaggcacgg gggaggggca aacaacagat ggctggcaac tagaaggcac
     6481 agtcgaggct gatcagcggg tttaaacggg ccctgctaga gattttccac actgactaaa
     6541 agggtctgag ggatctctag ttaccagagt cacacaacag acgggcacac actacttgaa
     6601 gcactcaagg caagctttat tgaggcttaa gcagtgggtt ccctagttag ccagagagct
     6661 cccaggctca gatctggtct aaccagagag acccagtaca agcaaaaagc agatcttgtc
     6721 ttcgttggga gtgaattagc ccttccagtc cccccttttc ttttaaaaag tggctaagat
     6781 ctacagctgc cttgtaagtc attggtctta aagtcgacgc ggggaggcgg cccaaaggga
     6841 gatccgactc gtctgagggc gaaggcgaag acgcggaaga ggccgcagag ccggcagcag
     6901 gccgcgggaa ggaaggtccg ctggattgag ggccgaaggg acgtagcaga aggacgtccc
     6961 gcgcagaatc caggtggcaa cacaggcgag cagccaagga aaggacgatg atttccccga
     7021 caacaccacg gaattgtcag tgcccaacag ccgagcccct gtccagcagc gggcaaggca
     7081 ggcggcgatg agttccgccg tggcaatagg gagggggaaa gcgaaagtcc cggaaaggag
     7141 ctgacaggtg gtggcaatgc cccaaccagt gggggttgcg tcagcaaaca cagtgcacac
     7201 cacgccacgt tgcctgacaa cgggccacaa ctcctcataa agagacagca accaggattt
     7261 atacaaggag gagaaaatga aagccatacg ggaagcaata gcatgataca aaggcattaa
     7321 agcagcgtat ccacatagcg taaaaggagc aacatagtta agaataccag tcaatctttc
     7381 acaaattttg taatccagag gttgattgtc gacttaacgc gtctatcggc gaagtacttc
     7441 tacacagcca tcggtccaga cggccgcgct tctgcgggcg atttgtgtac gcccgacagt
     7501 cccggctccg gatcggacga ttgcgtcgca tcgaccctgc gcccaagctg catcatcgaa
     7561 attgccgtca accaagctct gatagagttg gtcaagacca atgcggagca tatacgcccg
     7621 gagccgcggc gatcctgcaa gctccggatg cctccgctcg aagtagcgcg tctgctgctc
     7681 catacaagcc aaccacggcc tccagaagaa gatgttggcg acctcgtatt gggaatcccc
     7741 gaacatcgcc tcgctccagt caatgaccgc tgttatgcgg ccattgtccg tcaggacatt
     7801 gttggagccg aaatccgcgt gcacgaggtg ccggacttcg gggcagtcct cggcccaaag
     7861 catcagctca tcgagagcct gcgcgacgga cgcactgacg gtgtcgtcca tcacagtttg
     7921 ccagtgatac acatggggat cagcaatcgc gcatatgaaa tcacgccatg tagtgtattg
     7981 accgattcct tgcggtccga atgggccgaa cccgctcgtc tggctaagat cggccgcagc
     8041 gatcgcatcc atggcctccg cgaccggctg cagaacagcg ggcagttcgg tttcaggcag
     8101 gtcttgcaac gtgacaccct gtgcacggcg ggagatgcaa taggtcaggc tctcgctgaa
     8161 ttccccaatg tcaagcactt ccggaatcgg gagcgcggcc gatgcaaagt gccgataaac
     8221 ataacgatct ttgtagaaac catcggcgca gctatttacc cgcaggacat atccacgccc
     8281 tcctacatcg aagctgaaag cacgagattc ttcgccctcc gagagctgca tcaggtcgga
     8341 cacgctgtcg aacttttcga tcagaaactt ctcgacagac gtcgcggtga gttcaggctt
     8401 tttcggtcca ggattctctt cgacatctcc ggcttgtttc agcagagaga agtttgttgc
     8461 gccggatccc ttatcgtcat cgtctttgta atctttcttc ttcttagcct gtccagcctt
     8521 ctttgtggcg gcaggtcgct tgtcgcctcc cagctgagac aggtcgatcc gtgtctcgta
     8581 caggccggtg atgctctggt ggatcagggt ggcgtccagc acctctttgg tgctggtgta
     8641 cctcttccgg tcgatggtgg tgtcaaagta cttgaaggcg gcaggggctc ccagattggt
     8701 cagggtaaac aggtggatga tattctcggc ctgctctctg atgggcttat cccggtgctt
     8761 gttgtaggcg gacagcactt tgtccagatt agcgtcggcc aggatcactc tcttggagaa
     8821 ctcgctgatc tgctcgatga tctcgtccag gtagtgcttg tgctgttcca caaacagctg
     8881 tttctgctca ttatcctcgg gggagccctt cagcttctca tagtggctgg ccaggtacag
     8941 gaagttcaca tatttggagg gcagggccag ttcgtttccc ttctgcagtt cgccggcaga
     9001 ggccagcatt ctcttccggc cgttttccag ctcgaacagg gagtacttag gcagcttgat
     9061 gatcaggtcc tttttcactt ctttgtagcc cttggcttcc agaaagtcga tgggattctt
     9121 ctcgaagctg cttctttcca tgatggtgat ccccagcagc tctttcacac tcttcagttt
     9181 cttggacttg cccttttcca ctttggccac caccagcaca gaataggcca cggtggggct
     9241 gtcgaagccg ccgtacttct tagggtccca gtccttcttt ctggcgatca gcttatcgct
     9301 gttcctcttg ggcaggatag actctttgct gaagccgcct gtctgcacct cggtcttttt
     9361 cacgatattc acttggggca tgctcagcac tttccgcacg gtggcaaaat cccggccctt
     9421 atcccacacg atctccccgg tttcgccgtt tgtctcgatc agaggccgct tccggatctc
     9481 gccgttggcc agggtaatct cggtcttgaa aaagttcatg atgttgctgt agaagaagta
     9541 cttggcggta gccttgccga tttcctgctc gctcttggcg atcatcttcc gcacgtcgta
     9601 caccttgtag tcgccgtaca cgaactcgct ttccagctta gggtactttt tgatcagggc
     9661 ggttcccacg acggcgttca ggtaggcgtc gtgggcgtgg tggtagttgt tgatctcgcg
     9721 cactttgtaa aactggaaat ccttccggaa atcggacacc agcttggact tcagggtgat
     9781 cactttcact tcccggatca gcttgtcatt ctcgtcgtac ttagtgttca tccgggagtc
     9841 caggatctgt gccacgtgct ttgtgatctg ccgggtttcc accagctgtc tcttgatgaa
     9901 gccggcctta tccagttcgc tcaggccgcc tctctcggcc ttggtcagat tgtcgaactt
     9961 tctctgggta atcagcttgg cgttcagcag ctgccgccag tagttcttca tcttcttcac
    10021 gacctcttcg gagggcacgt tgtcgctctt gccccggttc ttgtcgcttc tggtcagcac
    10081 cttgttgtcg atggagtcgt ccttcagaaa gctctgaggc acgatatggt ccacatcgta
    10141 gtcggacagc cggttgatgt ccagttcctg gtccacgtac atatcccgcc cattctgcag
    10201 gtagtacagg tacagcttct cgttctgcag ctgggtgttt tccacggggt gttctttcag
    10261 gatctggctg cccagctctt tgatgccctc ttcgatccgc ttcattctct cgcggctgtt
    10321 cttctgtccc ttctgggtgg tctggttctc tctggccatt tcgatcacga tgttctcggg
    10381 cttgtgccgg cccatcactt tcacgagctc gtccaccacc ttcactgtct gcaggatgcc
    10441 cttcttaatg gcggggctgc cggccagatt ggcaatgtgc tcgtgcaggc tatcgccctg
    10501 gccggacacc tgggctttct ggatgtcctc tttaaaggtc aggctgtcgt cgtggatcag
    10561 ctgcatgaag tttctgttgg cgaagccgtc ggacttcagg aaatccagga ttgtcttgcc
    10621 ggactgcttg tcccggatgc cgttgatcag cttccggctc agcctgcccc agccggtgta
    10681 tctccgccgc ttcagctgct tcatcacttt gtcgtcgaac aggtgggcat aggttttcag
    10741 ccgttcctcg atcatctctc tgtcctcaaa cagtgtcagg gtcagcacga tatcttccag
    10801 aatgtcctcg ttttcctcat tgtccaggaa gtccttgtcc ttgataattt tcagcagatc
    10861 gtggtatgtg cccagggagg cgttgaaccg atcttccacg ccggagattt ccacggagtc
    10921 gaagcactcg attttcttga agtagtcctc tttcagctgc ttcacggtca ctttccggtt
    10981 ggtcttgaac agcaggtcca cgatggcctt tttctgctcg ccgctcagga aggcgggctt
    11041 tctcattccc tcggtcacgt atttcacttt ggtcagctcg ttatacacgg tgaagtactc
    11101 gtacagcagg ctgtgcttgg gcagcacctt ctcgttgggc aggttcttat cgaagttggt
    11161 catccgctcg atgaagctct gggcggaagc gcccttgtcc accacttcct cgaagttcca
    11221 gggggtgatg gtttcctcgc tctttctggt catccaggcg aatctgctgt ttcccctggc
    11281 cagagggccc acgtagtagg ggatgcggaa ggtcaggatc ttctcgatct tttcccggtt
    11341 gtccttcagg aatgggtaaa aatcttcctg ccgccgcaga atggcgtgca gctctcccag
    11401 gtggatctgg tgggggatgc tgccgttgtc gaaggtccgc tgcttccgca gcaggtcctc
    11461 tctgttcagc ttcacgagca gttcctcggt gccgtccatc ttttccagga tgggcttgat
    11521 gaacttgtag aactcttcct ggctggctcc gccgtcaatg tagccggcgt agccgttctt
    11581 gctctggtcg aagaaaatct ctttgtactt ctcaggcagc tgctgccgca cgagagcttt
    11641 cagcagggtc aggtcctggt ggtgctcgtc gtatctcttg atcatagagg cgctcagggg
    11701 ggccttggtg atctcggtgt tcactctcag gatgtcgctc agcaggatgg cgtcggacag
    11761 gttcttggcg gccagaaaca ggtcggcgta ctggtcgccg atctgggcca gcaggttgtc
    11821 caggtcgtcg tcgtaggtgt ccttgctcag ctgcagtttg gcatcctcgg ccaggtcgaa
    11881 gttgctcttg aagttggggg tcaggcccag gctcagggca atcaggtttc cgaacaggcc
    11941 attcttcttc tcgccgggca gctgggcgat cagattttcc agccgtctgc tcttgctcag
    12001 tctggcagac aggatggcct tggcgtccac gccgctggcg ttgatggggt tttcctcgaa
    12061 cagctggttg taggtctgca ccagctggat gaacagcttg tccacgtcgc tgttgtcggg
    12121 gttcaggtcg ccctcgatca ggaagtggcc ccggaacttg atcatgtggg ccagggccag
    12181 atagatcagc cgcaggtcgg ccttgtcggt gctgtccacc agtttctttc tcaggtggta
    12241 gatggtgggg tacttctcgt ggtaggccac ctcgtccacg atgttgccga agatggggtg
    12301 ccgctcgtgc ttcttatcct cttccaccag gaaggactct tccagtctgt ggaagaagct
    12361 gtcgtccacc ttggccatct cgttgctgaa gatctcttgc agatagcaga tccggttctt
    12421 ccgtctggtg tatcttcttc tggcggttct cttcagccgg gtggcctcgg ctgtttcgcc
    12481 gctgtcgaac agcagggctc cgatcaggtt cttcttgatg ctgtgccggt cggtgttgcc
    12541 cagcaccttg aatttcttgc tgggcacctt gtactcgtcg gtgatcacgg cccagcccac
    12601 agagttggtg ccgatgtcca ggccgatgct gtacttcttg tccatggtgg cagcgctcta
    12661 gaaccggtcc tgtgttctgg cggcaaaccc gttgcgaaaa agaacgttca cggcgactac
    12721 tgcacttata tacggttctc ccccaccctc gggaaaaagg cggagccagt acacgacatc
    12781 actttcccag tttaccccgc gccaccttct ctaggcaccg gatcaattgc cgacccctcc
    12841 ccccaacttc tcggggactg tgggcgatgt gcgctctgcc cactgacggg caccggagcc
    12901 aattcccact cctttcaaga cctagctagc gaattcaaaa aagcaccgac tcggtgccac
    12961 tttttcaagt tgataacgga ctagccttat tttaacttgc tatttctagc tctaaaaccg
    13021 gaacgtatag gacatacggg tgtttcgtcc tttccacaag atatataaag ccaagaaatc
    13081 gaaatacttt caagttacgg taagcatatg atagtccatt ttaaaacata attttaaaac
    13141 tgcaaactac ccaagaaatt attactttct acgtcacgta ttttgtacta atatctttgt
    13201 gtttacagtc aaattaattc taattatctc tctaacagcc ttgtatcgta tatgcaaata
    13261 tgaaggaatc atgggaaata ggccctcggt accttaatta accaaactgg atctctgctg
    13321 tccctgtaat aaacccgaaa attttgaatt tttgtaattt gtttttgtaa ttctttagtt
    13381 tgtatgtctg ttgctattat
//
