LOCUS       sequence-466836-       12824 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  knockout Crls.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12824)
  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 12824)
  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-sgCrls1-2#
FEATURES             Location/Qualifiers
     source          1..12824
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(69..113)
                     /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    298..531
                     /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    1024..1149
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             1196..1376
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     promoter        complement(1394..1592)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(1419..1439)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        1594..1973
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     2145..2164
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        2239..2343
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2344..2412,2413..3204)
                     /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(2562..2581)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3375..3963
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3864..3883
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4117..4134
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4251..4272
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4287..4292,4293..4310,4311..4317)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4325..4341
                     /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     4330..4352
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4349..4365
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4349..4365
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    4414..4535
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4425..4444)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4479..4498
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(4665..5039)
                     /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(5058..5105)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(5153..5482)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5167..5302
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(5221..5240)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     5467..5487
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(5496..5924)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5606..5627)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5818..5837
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    5970..6194
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     6183..6200
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     LTR             6226..6459
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     misc_feature    6531..7119
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             6637..6648
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     7046..7066
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(7144..7542)
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     CDS             complement(7543..7599)
                     /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(7609..7632)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             complement(7633..7680)
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             complement(7681..11784)
                     /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(7681..11784)
                     /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      11781..11790
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(11809..12020)
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     misc_RNA        complement(12082..12157)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(12179..12427)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(12237..12256)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(12407..12427)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     misc_feature    12478..12595
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
ORIGIN
        1 aagcctccta ctatcattat gaataatttt atataccaca gccaatttgt tatgttaaac
       61 caattccaca aacttgccca tttatctaat tccaataatt cttgttcatt cttttcttgc
      121 tggttttgcg attcttcaat taaggagtgt attaagcttg tgtaattgtt aatttctctg
      181 tcccactcca tccaggtcgt gtgattccaa atctgttcca gagatttatt actccaacta
      241 gcattccaag gcacagcagt ggtgcaaatg agttttccag agcaacccca aatccccagg
      301 agctgttgat cctttaggta tctttccaca gccaggattc ttgcctggag ctgcttgatg
      361 ccccagactg tgagttgcaa cagatgctgt tgcgcctcaa tagccctcag caaattgttc
      421 tgctgctgca ctataccaga caataattgt ctggcctgta ccgtcagcgt cattgacgct
      481 gcgcccatag tgcttcctgc tgctcccaag aacccaagga acaaagctcc tattcccact
      541 gctctttttt ctctctgcac cactcttctc tttgccttgg tgggtgctac tcctaatggt
      601 tcaattttta ctactttata tttatataat tcacttctcc aattgtccct catatctcct
      661 cctccaggtc tgaagatcag cggccgcttg ctgtgcggtg gtcttacttt tgttttgctc
      721 ttcctctatc ttgtctaaag cttccttggt gtcttttatc tctatccttt gatgcacaca
      781 atagagggtt gctactgtat tatataatga tctaagttct tctgatcctg tctgaaggga
      841 tggttgtagc tgtcccagta tttgtctaca gccttctgat gtttctaaca ggccaggatt
      901 aactgcgaat cgttctagct ccctgcttgc ccatactata tgttttaatt tatatttttt
      961 ctttccccct ggccttaacc gaattttttc ccatcgcgat ctaattctcc cccgcttaat
     1021 actgacgctc tcgcacccat ctctctcctt ctagcctccg ctagtcaaaa tttttggcgt
     1081 actcaccagt cgccgcccct cgcctcttgc cgtgcgcgct tcagcaagcc gagtcctgcg
     1141 tcgagagagc tcctctggtt tccctttcgc tttcaagtcc ctgttcgggc gccactgcta
     1201 gagattttcc acactgacta aaagggtctg agggatctct agttaccaga gtcacacaac
     1261 agacgggcac acactacttg aagcactcaa ggcaagcttt attgaggctt aagcagtggg
     1321 ttccctagtt agccagagag ctcccaggct cagatctggt ctaaccagag agacccagta
     1381 caggcaaaac gcgctgctta tatagacctc ccaccgtaca cgcctaccgc ccatttgcgt
     1441 caatggggcg gagttgttac gacattttgg aaagtcccgt tgattttggt gccaaaacaa
     1501 actcccattg acgtcaatgg ggtggagact tggaaatccc cgtgagtcaa accgctatcc
     1561 acgcccattg atgtactgcc aaaaccgcat caccatggta atagcgatga ctaatacgta
     1621 gatgtactgc caagtaggaa agtcccataa ggtcatgtac tgggcataat gccaggcggg
     1681 ccatttaccg tcattgacgt caataggggg cgtacttggc atatgataca cttgatgtac
     1741 tgccaagtgg gcagtttacc gtaaatactc cacccattga cgtcaatgga aagtccctat
     1801 tggcgttact atgggaacat acgtcattat tgacgtcaat gggcgggggt cgttgggcgg
     1861 tcagccaggc gggccattta ccgtaagtta tgtaacgcgg aactccatat atgggctatg
     1921 aactaatgac cccgtaattg attactatta ataactagtc aataatcaat gtcaacgcgt
     1981 atatctggcc cgtacatcgc gaagcagcgc aaaacgccta accctaagca gattcttcat
     2041 gcaattgtcg gtcaagcctt gccttgttgt agcttaaatt ttgctcgcgc actactcagc
     2101 gacctccaac acacaagcag ggagcagata ctggcttaac tatgcggcat cagagcagat
     2161 tgtactgaga gtgcaccata ggggatcggg agatctcccg atccgtcgac gtcaggtggc
     2221 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     2281 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     2341 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
     2401 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
     2461 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
     2521 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
     2581 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
     2641 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
     2701 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
     2761 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
     2821 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
     2881 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
     2941 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
     3001 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
     3061 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
     3121 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
     3181 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
     3241 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     3301 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     3361 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     3421 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     3481 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     3541 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     3601 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     3661 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     3721 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     3781 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     3841 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     3901 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     3961 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     4021 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     4081 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
     4141 gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc
     4201 tggcacgaca ggtttcccga ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt
     4261 tagctcactc attaggcacc ccaggcttta cactttatgc ttccggctcg tatgttgtgt
     4321 ggaattgtga gcggataaca atttcacaca ggaaacagct atgaccatga ttacgccaag
     4381 ctctagctag aggtcgacgg tatacagaca tgataagata cattgatgag tttggacaaa
     4441 ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat gctattgctt
     4501 tatttgtaac cattataagc tgcaataaac aagttggggt gggcgaagaa ctccagcatg
     4561 agatccccgc gctggaggat catccagccg gcgtcccgga aaacgattcc gaagcccaac
     4621 ctttcataga aggcggcggt ggaatcgaaa tctcgtagca cgtgtcagtc ctgctcctcg
     4681 gccacgaagt gcacgcagtt gccggccggg tcgcgcaggg cgaactcccg cccccacggc
     4741 tgctcgccga tctcggtcat ggccggcccg gaggcgtccc ggaagttcgt ggacacgacc
     4801 tccgaccact cggcgtacag ctcgtccagg ccgcgcaccc acacccaggc cagggtgttg
     4861 tccggcacca cctggtcctg gaccgcgctg atgaacaggg tcacgtcgtc ccggaccaca
     4921 ccggcgaagt cgtcctccac gaagtcccgg gagaacccga gccggtcggt ccagaactcg
     4981 accgctccgg cgacgtcgcg cgcggtgagc accggaacgg cactggtcaa cttggccatg
     5041 gtttagttcc tcaccttgtc gtattatact atgccgatat actatgccga tgattaattg
     5101 tcaacacgtg ctgatcagat ccgaaaatgg atatacaagc tcccgggagc tttttgcaaa
     5161 agcctaggcc tccaaaaaag cctcctcact acttctggaa tagctcagag gcagaggcgg
     5221 cctcggcctc tgcataaata aaaaaaatta gtcagccatg gggcggagaa tgggcggaac
     5281 tgggcggagt taggggcggg atgggcggag ttaggggcgg gactatggtt gctgactaat
     5341 tgagatgcat gctttgcata cttctgcctg ctggggagcc tggggacttt ccacacctgg
     5401 ttgctgacta attgagatgc atgctttgca tacttctgcc tgctggggag cctggggact
     5461 ttccacaccc taactgacac acattccaca gaattaattc gcgttaaatt tttgttaaat
     5521 cagctcattt tttaaccaat aggccgaaat cggcaaaatc ccttataaat caaaagaata
     5581 gaccgagata gggttgagtg ttgttccagt ttggaacaag agtccactat taaagaacgt
     5641 ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac tacgtgaacc
     5701 atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc ggaaccctaa
     5761 agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga gaaaggaagg
     5821 gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt gtagcggtca cgctgcgcgt
     5881 aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtggggat accccctaga
     5941 gccccagctg gttctttccg cctcagaagc catagagccc accgcatccc cagcatgcct
     6001 gctattgtct tcccaatcct cccccttgct gtcctgcccc accccacccc ccagaataga
     6061 atgacaccta ctcagacaat gcgatgcaat ttcctcattt tattaggaaa ggacagtggg
     6121 agtggcacct tccagggtca aggaaggcac gggggagggg caaacaacag atggctggca
     6181 actagaaggc acagtcgagg ctgatcagcg ggtttaaacg ggccctgcta gagattttcc
     6241 acactgacta aaagggtctg agggatctct agttaccaga gtcacacaac agacgggcac
     6301 acactacttg aagcactcaa ggcaagcttt attgaggctt aagcagtggg ttccctagtt
     6361 agccagagag ctcccaggct cagatctggt ctaaccagag agacccagta caagcaaaaa
     6421 gcagatcttg tcttcgttgg gagtgaatta gcccttccag tccccccttt tcttttaaaa
     6481 agtggctaag atctacagct gccttgtaag tcattggtct taaagtcgac gcggggaggc
     6541 ggcccaaagg gagatccgac tcgtctgagg gcgaaggcga agacgcggaa gaggccgcag
     6601 agccggcagc aggccgcggg aaggaaggtc cgctggattg agggccgaag ggacgtagca
     6661 gaaggacgtc ccgcgcagaa tccaggtggc aacacaggcg agcagccaag gaaaggacga
     6721 tgatttcccc gacaacacca cggaattgtc agtgcccaac agccgagccc ctgtccagca
     6781 gcgggcaagg caggcggcga tgagttccgc cgtggcaata gggaggggga aagcgaaagt
     6841 cccggaaagg agctgacagg tggtggcaat gccccaacca gtgggggttg cgtcagcaaa
     6901 cacagtgcac accacgccac gttgcctgac aacgggccac aactcctcat aaagagacag
     6961 caaccaggat ttatacaagg aggagaaaat gaaagccata cgggaagcaa tagcatgata
     7021 caaaggcatt aaagcagcgt atccacatag cgtaaaagga gcaacatagt taagaatacc
     7081 agtcaatctt tcacaaattt tgtaatccag aggttgatta ccgataagct tgatatcgaa
     7141 ttcttagccc tcccacacat aaccagaggg cagcaattca cgaatcccaa ctgccgtcgg
     7201 ctgtccatca ctgtccttca ctatggcttt gatcccagga tgcagatcga gaagcacctg
     7261 tcggcaccgt ccgcaggggc tcaagatgcc cctgttctca tttccgatcg cgacgataca
     7321 agtcaggttg ccagctgccg cagcagcagc agtgcccagc accacgagtt ctgcacaagg
     7381 tcccccagta aaatgatata cattgacacc agtgaagatg cggccgtcgc tagagagagc
     7441 tgcgctggcg acgctgtagt cttcagagat ggggatgctg ttgattgtag ccgttgctct
     7501 ttcaatgagg gtggattctt cttgagacaa aggcttggcc atcggtccag gattctcttc
     7561 gacatctccg gcttgtttca gcagagagaa gtttgttgcg ccggatccct tatcgtcatc
     7621 gtctttgtaa tctttcttct tcttagcctg tccagccttc tttgtggcgg caggtcgctt
     7681 gtcgcctccc agctgagaca ggtcgatccg tgtctcgtac aggccggtga tgctctggtg
     7741 gatcagggtg gcgtccagca cctctttggt gctggtgtac ctcttccggt cgatggtggt
     7801 gtcaaagtac ttgaaggcgg caggggctcc cagattggtc agggtaaaca ggtggatgat
     7861 attctcggcc tgctctctga tgggcttatc ccggtgcttg ttgtaggcgg acagcacttt
     7921 gtccagatta gcgtcggcca ggatcactct cttggagaac tcgctgatct gctcgatgat
     7981 ctcgtccagg tagtgcttgt gctgttccac aaacagctgt ttctgctcat tatcctcggg
     8041 ggagcccttc agcttctcat agtggctggc caggtacagg aagttcacat atttggaggg
     8101 cagggccagt tcgtttccct tctgcagttc gccggcagag gccagcattc tcttccggcc
     8161 gttttccagc tcgaacaggg agtacttagg cagcttgatg atcaggtcct ttttcacttc
     8221 tttgtagccc ttggcttcca gaaagtcgat gggattcttc tcgaagctgc ttctttccat
     8281 gatggtgatc cccagcagct ctttcacact cttcagtttc ttggacttgc ccttttccac
     8341 tttggccacc accagcacag aataggccac ggtggggctg tcgaagccgc cgtacttctt
     8401 agggtcccag tccttctttc tggcgatcag cttatcgctg ttcctcttgg gcaggataga
     8461 ctctttgctg aagccgcctg tctgcacctc ggtctttttc acgatattca cttggggcat
     8521 gctcagcact ttccgcacgg tggcaaaatc ccggccctta tcccacacga tctccccggt
     8581 ttcgccgttt gtctcgatca gaggccgctt ccggatctcg ccgttggcca gggtaatctc
     8641 ggtcttgaaa aagttcatga tgttgctgta gaagaagtac ttggcggtag ccttgccgat
     8701 ttcctgctcg ctcttggcga tcatcttccg cacgtcgtac accttgtagt cgccgtacac
     8761 gaactcgctt tccagcttag ggtacttttt gatcagggcg gttcccacga cggcgttcag
     8821 gtaggcgtcg tgggcgtggt ggtagttgtt gatctcgcgc actttgtaaa actggaaatc
     8881 cttccggaaa tcggacacca gcttggactt cagggtgatc actttcactt cccggatcag
     8941 cttgtcattc tcgtcgtact tagtgttcat ccgggagtcc aggatctgtg ccacgtgctt
     9001 tgtgatctgc cgggtttcca ccagctgtct cttgatgaag ccggccttat ccagttcgct
     9061 caggccgcct ctctcggcct tggtcagatt gtcgaacttt ctctgggtaa tcagcttggc
     9121 gttcagcagc tgccgccagt agttcttcat cttcttcacg acctcttcgg agggcacgtt
     9181 gtcgctcttg ccccggttct tgtcgcttct ggtcagcacc ttgttgtcga tggagtcgtc
     9241 cttcagaaag ctctgaggca cgatatggtc cacatcgtag tcggacagcc ggttgatgtc
     9301 cagttcctgg tccacgtaca tatcccgccc attctgcagg tagtacaggt acagcttctc
     9361 gttctgcagc tgggtgtttt ccacggggtg ttctttcagg atctggctgc ccagctcttt
     9421 gatgccctct tcgatccgct tcattctctc gcggctgttc ttctgtccct tctgggtggt
     9481 ctggttctct ctggccattt cgatcacgat gttctcgggc ttgtgccggc ccatcacttt
     9541 cacgagctcg tccaccacct tcactgtctg caggatgccc ttcttaatgg cggggctgcc
     9601 ggccagattg gcaatgtgct cgtgcaggct atcgccctgg ccggacacct gggctttctg
     9661 gatgtcctct ttaaaggtca ggctgtcgtc gtggatcagc tgcatgaagt ttctgttggc
     9721 gaagccgtcg gacttcagga aatccaggat tgtcttgccg gactgcttgt cccggatgcc
     9781 gttgatcagc ttccggctca gcctgcccca gccggtgtat ctccgccgct tcagctgctt
     9841 catcactttg tcgtcgaaca ggtgggcata ggttttcagc cgttcctcga tcatctctct
     9901 gtcctcaaac agtgtcaggg tcagcacgat atcttccaga atgtcctcgt tttcctcatt
     9961 gtccaggaag tccttgtcct tgataatttt cagcagatcg tggtatgtgc ccagggaggc
    10021 gttgaaccga tcttccacgc cggagatttc cacggagtcg aagcactcga ttttcttgaa
    10081 gtagtcctct ttcagctgct tcacggtcac tttccggttg gtcttgaaca gcaggtccac
    10141 gatggccttt ttctgctcgc cgctcaggaa ggcgggcttt ctcattccct cggtcacgta
    10201 tttcactttg gtcagctcgt tatacacggt gaagtactcg tacagcaggc tgtgcttggg
    10261 cagcaccttc tcgttgggca ggttcttatc gaagttggtc atccgctcga tgaagctctg
    10321 ggcggaagcg cccttgtcca ccacttcctc gaagttccag ggggtgatgg tttcctcgct
    10381 ctttctggtc atccaggcga atctgctgtt tcccctggcc agagggccca cgtagtaggg
    10441 gatgcggaag gtcaggatct tctcgatctt ttcccggttg tccttcagga atgggtaaaa
    10501 atcttcctgc cgccgcagaa tggcgtgcag ctctcccagg tggatctggt gggggatgct
    10561 gccgttgtcg aaggtccgct gcttccgcag caggtcctct ctgttcagct tcacgagcag
    10621 ttcctcggtg ccgtccatct tttccaggat gggcttgatg aacttgtaga actcttcctg
    10681 gctggctccg ccgtcaatgt agccggcgta gccgttcttg ctctggtcga agaaaatctc
    10741 tttgtacttc tcaggcagct gctgccgcac gagagctttc agcagggtca ggtcctggtg
    10801 gtgctcgtcg tatctcttga tcatagaggc gctcaggggg gccttggtga tctcggtgtt
    10861 cactctcagg atgtcgctca gcaggatggc gtcggacagg ttcttggcgg ccagaaacag
    10921 gtcggcgtac tggtcgccga tctgggccag caggttgtcc aggtcgtcgt cgtaggtgtc
    10981 cttgctcagc tgcagtttgg catcctcggc caggtcgaag ttgctcttga agttgggggt
    11041 caggcccagg ctcagggcaa tcaggtttcc gaacaggcca ttcttcttct cgccgggcag
    11101 ctgggcgatc agattttcca gccgtctgct cttgctcagt ctggcagaca ggatggcctt
    11161 ggcgtccacg ccgctggcgt tgatggggtt ttcctcgaac agctggttgt aggtctgcac
    11221 cagctggatg aacagcttgt ccacgtcgct gttgtcgggg ttcaggtcgc cctcgatcag
    11281 gaagtggccc cggaacttga tcatgtgggc cagggccaga tagatcagcc gcaggtcggc
    11341 cttgtcggtg ctgtccacca gtttctttct caggtggtag atggtggggt acttctcgtg
    11401 gtaggccacc tcgtccacga tgttgccgaa gatggggtgc cgctcgtgct tcttatcctc
    11461 ttccaccagg aaggactctt ccagtctgtg gaagaagctg tcgtccacct tggccatctc
    11521 gttgctgaag atctcttgca gatagcagat ccggttcttc cgtctggtgt atcttcttct
    11581 ggcggttctc ttcagccggg tggcctcggc tgtttcgccg ctgtcgaaca gcagggctcc
    11641 gatcaggttc ttcttgatgc tgtgccggtc ggtgttgccc agcaccttga atttcttgct
    11701 gggcaccttg tactcgtcgg tgatcacggc ccagcccaca gagttggtgc cgatgtccag
    11761 gccgatgctg tacttcttgt ccatggtggc agcgctctag aaccggtcct gtgttctggc
    11821 ggcaaacccg ttgcgaaaaa gaacgttcac ggcgactact gcacttatat acggttctcc
    11881 cccaccctcg ggaaaaaggc ggagccagta cacgacatca ctttcccagt ttaccccgcg
    11941 ccaccttctc taggcaccgg atcaattgcc gacccctccc cccaacttct cggggactgt
    12001 gggcgatgtg cgctctgccc actgacgggc accggagcca attcccactc ctttcaagac
    12061 ctagctagcg aattcaaaaa agcaccgact cggtgccact ttttcaagtt gataacggac
    12121 tagccttatt ttaacttgct atttctagct ctaaaacgtt tcgagcaata aatccatcgg
    12181 tgtttcgtcc tttccacaag atatataaag ccaagaaatc gaaatacttt caagttacgg
    12241 taagcatatg atagtccatt ttaaaacata attttaaaac tgcaaactac ccaagaaatt
    12301 attactttct acgtcacgta ttttgtacta atatctttgt gtttacagtc aaattaattc
    12361 taattatctc tctaacagcc ttgtatcgta tatgcaaata tgaaggaatc atgggaaata
    12421 ggccctcggt accttaatta accaaactgg atctctgctg tccctgtaat aaacccgaaa
    12481 attttgaatt tttgtaattt gtttttgtaa ttctttagtt tgtatgtctg ttgctattat
    12541 gtctactatt ctttcccctg cactgtaccc cccaatcccc ccttttcttt taaaattgtg
    12601 gatgaatact gccatttgtc tgcagaattg gcgcacgcag tgccgatccg ttcactaatc
    12661 gaatggatct gtctctgtct ctctctccac cttcttcttc tattccttcg ggcctgtcgg
    12721 gtcccctcgg ggttgggagg tgggtctgaa acgataatgg tgaatatccc tgcctaactc
    12781 tattcactat agaaagtaca gcaaaaacta ttcttaaacc tacc
//
