LOCUS       sequence-412251-       12286 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Inducibly expresses human Caspase 8 fused to light-activatable Cry2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12286)
  AUTHORS   Shkarina K, Hasel de Carvalho E, Santos JC, Ramos S, Leptin M, Broz 
            P
  TITLE     Optogenetic activators of apoptosis, necroptosis, and pyroptosis.
  JOURNAL   J Cell Biol. 2022 Jun 6;221(6). pii: 213145. doi: 
            10.1083/jcb.202109038. Epub 2022 Apr 14.
  PUBMED    35420640
REFERENCE   2  (bases 1 to 12286)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Cell 
            Biol."; date: "2022-06-6"; volume: "221(6). pii: 213145. doi"; 
            pages: " 10.1083/jcb.202109038. Epub 2022 Apr 14"
COMMENT     Sequence Label: pLVX-opto-hCaspase-8
FEATURES             Location/Qualifiers
     source          1..12286
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /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     956..978
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     975..991
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     LTR             1119..1752
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    1959..2547
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             2065..2076
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     2474..2494
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2561..3160)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(2644..2664)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     3141..3160
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(3169..3498)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3183..3318
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             complement(3509..4255)
                     /codon_start=1
                     /product="modified rtTA protein that binds tightly to 
                     promoters containing the tet operator in the presence of 
                     doxycycline"
                     /label=Tet-On(R) 3G
                     /translation="MSRLDKSKVINSALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHSCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLKQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     promoter        complement(4274..4784)
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     complement(4337..4355)
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     promoter        4801..5168
                     /label=TRE3GS promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G, modified to eliminate
                     binding sites for endogenous mammalian transcription 
                     factors"
     protein_bind    4809..4827
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4845..4863
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4881..4899
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4917..4935
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4953..4971
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    4989..5007
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    5025..5043
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             5174..5881
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLRDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(5319..5337)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     5587..5606
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             5897..7390
                     /codon_start=1
                     /product="N-terminal photolyase homology region of 
                     Arabidopsis thaliana cryptochrome 2"
                     /label=CRY2PHR
                     /note="binds to CIB1 upon exposure to blue light (Kennedy 
                     et al., 2010)"
                     /translation="MKMDKKTIVWFRRDLRIEDNPALAAAAHEGSVFPVFIWCPEEEGQ
                     FYPGRASRWWMKQSLAHLSQSLKALGSDLTLIKTHNTISAILDCIRVTGATKVVFNHLY
                     DPVSLVRDHTVKEKLVERGISVQSYNGDLLYEPWEIYCEKGKPFTSFNSYWKKCLDMSI
                     ESVMLPPPWRLMPITAAAEAIWACSIEELGLENEAEKPSNALLTRAWSPGWSNADKLLN
                     EFIEKQLIDYAKNSKKVVGNSTSLLSPYLHFGEISVRHVFQCARMKQIIWARDKNSEGE
                     ESADLFLRGIGLREYSRYICFNFPFTHEQSLLSHLRFFPWDADVDKFKAWRQGRTGYPL
                     VDAGMRELWATGWMHNRIRVIVSSFAVKFLLLPWKWGMKYFWDTLLDADLECDILGWQY
                     ISGSIPDGHELDRLDNPALQGAKYDPEGEYIRQWLPELARLPTEWIHHPWDAPLTVLKA
                     SGVELGTNYAKPIVDIDTARELLAKAISRTRGAQIMIGAA"
     polyA_signal    8481..8615
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    8659..8774
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1 (lacking the first T)"
     CDS             complement(9037..9081)
                     /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    9266..9499
                     /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    9996..10121
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             10168..10801
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     polyA_signal    10969..11103
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        11151..11255
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(11256..11324,11325..12116)
                     /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(11474..11493)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga ccatgattac gccaagctct agctagaggt
     1021 cgacggtaaa cgctagcaat gtcaaggcct ctcactctct gatattcatt tctttgcaag
     1081 ttataaatac tgaataataa gatgacatga actactactg ctagagattt tccacactga
     1141 ctaaaagggt ctgagggatc tctagttacc agagtcacac aacagacggg cacacactac
     1201 ttgaagcact caaggcaagc tttattgagg cttaagcagt gggttcccta gttagccaga
     1261 gagctcccag gctcagatct ggtctaacca gagagaccca gtacaggcaa aaagcagctg
     1321 cttatatgca ggatctgagg gctcgccact ccccagtccc gcccaggcca cgcctccctg
     1381 gaaagtcccc agcggaaagt cccttgtagc aagctcgata tcagcagttc ttgaagtact
     1441 ccggatgcag ctctcgggcc acgtgatgaa atgctaggcg gctgtcaaac ctccactcta
     1501 acacttctct ctccgggtca tccatcccat gcaggctcac agggtgtaac aagctggtgt
     1561 tctctccttt attggcctct tctaccttat ctggctcaac tggtactagc ttgtagcacc
     1621 atccaaaggt cagtggatat ctgacccctg gccctggtgt gtagttctgc taatcaggga
     1681 agtagccttg tgtgtggtag atccacagat caaggatatc ttgtcttcgt tgggagtgaa
     1741 ttagcccttc cagtcccctc ttttctttta aaaagtggct aagatctaca gctgccttgt
     1801 aagtcattgg tcttaaaggt acctgaggtg tgactggaaa acccacctcc tcctcctctt
     1861 gtgcttctag ccaggcacaa tcagcattgg tagctgctgt attgctactt gtgattgctc
     1921 catgtttttc taggtctcga ctgcagaatt aattccaggc ggggaggcgg cccaaaggga
     1981 gatccgactc gtctgagggc gaaggcgaag acgcggaaga ggccgcagag ccggcagcag
     2041 gccgcgggaa ggaaggtccg ctggattgag ggccgaaggg acgtagcaga aggacgtccc
     2101 gcgcagaatc caggtggcaa cacaggcgag cagccatgga aaggacgtca gcttccccga
     2161 caacaccacg gaattgtcag tgcccaacag ccgagcccct gtccagcagc gggcaaggca
     2221 ggcggcgatg agttccgccg tggcaatagg gagggggaaa gcgaaagtcc cggaaaggag
     2281 ctgacaggtg gtggcaatgc cccaaccagt gggggttgcg tcagcaaaca cagtgcacac
     2341 cacgccacgt tgcctgacaa cgggccacaa ctcctcataa agagacagca accaggattt
     2401 atacaaggag gagaaaatga aagccatacg ggaagcaata gcatgataca aaggcattaa
     2461 agcagcgtat ccacatagcg taaaaggagc aacatagtta agaataccag tcaatctttc
     2521 acaaattttg taatccagag gttgattgtt ccagacgcgt tcaggcaccg ggcttgcggg
     2581 tcatgcacca ggtgcgcggt ccttcgggca cctcgacgtc ggcggtgacg gtgaagccga
     2641 gccgctcgta gaaggggagg ttgcggggcg cggaggtctc caggaaggcg ggcaccccgg
     2701 cgcgctcggc cgcctccact ccggggagca cgacggcgct gcccagaccc ttgccctggt
     2761 ggtcgggcga gacgccgacg gtggccagga accacgcggg ctccttgggc cggtgcggcg
     2821 ccaggaggcc ttccatctgt tgctgcgcgg ccagccggga accgctcaac tcggccatgc
     2881 gcgggccgat ctcggcgaac accgcccccg cttcgacgct ctccggcgtg gtccagaccg
     2941 ccaccgcggc gccgtcgtcc gcgacccaca ccttgccgat gtcgagcccg acgcgcgtga
     3001 ggaagagttc ttgcagctcg gtgacccgct cgatgtggcg gtccgggtcg acggtgtggc
     3061 gcgtggcggg gtagtcggcg aacgcggcgg cgagggtgcg tacggcccgg gggacgtcgt
     3121 cgcgggtggc gaggcgcacc gtgggcttgt actcggtcat ggtcgcgttt tgcaaaagcc
     3181 taggcctcca aaaaagcctc ctcactactt ctggaatagc tcagaggccg aggcggcctc
     3241 ggcctctgca taaataaaaa aaattagtca gccatggggc ggagaatggg cggaactggg
     3301 cggagttagg ggcgggatgg gcggagttag gggcgggact atggttgctg actaattgag
     3361 atgcatgctt tgcatacttc tgcctgctgg ggagcctggg gactttccac acctggttgc
     3421 tgactaattg agatgcatgc tttgcatact tctgcctgct ggggagcctg gggactttcc
     3481 acaccctaac tgacacacat tcgcgcgttt acccggggag catgtcaagg tcaaaatcgt
     3541 caagagcgtc agcaggcagc atatcaaggt caaagtcgtc aagggcatcg gctgggagca
     3601 tgtctaagtc aaaatcgtca agggcgtcgg tcggcccgcc gctttcgcac tttagctgtt
     3661 tctccaggcc acatatgatt agttccaggc cgaaaaggaa ggcaggttcg gctccctgcc
     3721 ggtcgaacag ctcaattgct tgtttcagaa gtgggggcat agaatcggtg gtaggtgtct
     3781 ctctttcctc ttttgctact tgatgctcct gttcctccaa tacgcagccc agtgtaaagt
     3841 ggcccacggc ggacagagcg tacagtgcgt tctccaggga gaagccttgc tgacacagga
     3901 acgcgagctg attttccagg gtttcgtact gtttctctgt tgggcgggtg ccgagatgca
     3961 ctttagcccc gtcgcgatgt gagaggagag cacagcggta tgacttggcg ttgttccgca
     4021 gaaagtcttg ccatgactcg ccttccaggg ggcaggagtg ggtatgatgc ctgtccagca
     4081 tctcgattgg cagggcatcg agcagggccc gcttgttctt cacgtgccag tacagggtag
     4141 gctgctcaac tcccagcttt tgagcgagtt tccttgtcgt caggccttcg ataccgactc
     4201 cattgagtaa ttccagagca gagtttatga ctttgctctt gtccagtcta gacatggtaa
     4261 ttcgatgatc cccctgggga gagaggtcgg tgattcggtc aacgagggag ccgactgccg
     4321 acgtgcgctc cggaggcttg cagaatgcgg aacaccgcgc gggcaggaac agggcccaca
     4381 ctaccgcccc acaccccgcc tcccgcaccg ccccttcccg gccgctgctc tcggcgcgcc
     4441 ctgctgagca gccgctattg gccacagccc atcgcggtcg gcgcgctgcc attgctccct
     4501 ggcgctgtcc gtctgcgagg gtactagtga gacgtgcggc ttccgtttgt cacgtccggc
     4561 acgccgcgaa ccgcaaggaa ccttcccgac ttaggggcgg agcaggaagc gtcgccgggg
     4621 ggcccacaag ggtagcggcg aagatccggg tgacgctgcg aacggacgtg aagaatgtgc
     4681 gagacccagg gtcggcgccg ctgcgtttcc cggaaccacg cccagagcag ccgcgtccct
     4741 gcgcaaaccc agggctgcct tggaaaaggc gcaaccccaa ccccgtggaa ttatcacctc
     4801 gagtttactc cctatcagtg atagagaacg tatgaagagt ttactcccta tcagtgatag
     4861 agaacgtatg cagactttac tccctatcag tgatagagaa cgtataagga gtttactccc
     4921 tatcagtgat agagaacgta tgaccagttt actccctatc agtgatagag aacgtatcta
     4981 cagtttactc cctatcagtg atagagaacg tatatccagt ttactcccta tcagtgatag
     5041 agaacgtata agctttgctt atgtaaacca gggcgcctat aaaagagtgc tgattttttg
     5101 agtaaacttc aattccacaa cacttttgtc ttataccaac tttccgtacc acttcctacc
     5161 ctcgtaaaga attatggtga gcaagggcga ggaggataac atggccatca tcaaggagtt
     5221 catgcgcttc aaggtgcaca tggagggctc cgtgaacggc cacgagttcg agatcgaggg
     5281 cgagggcgag ggccgcccat acgagggcac ccagaccgcc aagctgaagg tgaccaaggg
     5341 tggccccctg cccttcgcct gggacatcct gtcccctcag ttcatgtacg gctccaaggc
     5401 ctacgtgaag caccccgccg acatccccga ctacttgaag ctgtccttcc ccgagggctt
     5461 caagtgggag cgcgtgatga acttcgagga cggcggcgtg gtgaccgtga cccaggactc
     5521 ctccctgcag gacggcgagt tcatctacaa ggtgaagctg cgcggcacca acttcccctc
     5581 cgacggcccc gtaatgcaga agaagaccat gggctgggag gcctcctccg agcggatgta
     5641 ccccgaggac ggcgccctga agggcgagat caagcagagg ctgaagctga gggacggcgg
     5701 ccactacgac gctgaggtca agaccaccta caaggccaag aagcccgtgc agctgcccgg
     5761 cgcctacaac gtcaacatca agttggacat cacctcccac aacgaggact acaccatcgt
     5821 ggaacagtac gaacgcgccg agggccgcca ctccaccggc ggcatggacg agctgtacaa
     5881 gggaggagga ggaatgatga agatggacaa aaagactata gtttggttta gaagagacct
     5941 aaggattgag gataatcctg cattagcagc agctgctcac gaaggatctg tttttcctgt
     6001 cttcatttgg tgtcctgaag aagaaggaca gttttatcct ggaagagctt caagatggtg
     6061 gatgaaacaa tcacttgctc acttatctca atccttgaag gctcttggat ctgacctcac
     6121 tttaatcaaa acccacaaca cgatttcagc gatcttggat tgtatccgcg ttaccggtgc
     6181 tacaaaagtc gtctttaacc acctctatga tcctgtttcg ttagttcggg accataccgt
     6241 aaaggagaag ctggtggaac gtgggatctc tgtgcaaagc tacaatggag atctattgta
     6301 tgaaccgtgg gagatatact gcgaaaaggg caaacctttt acgagtttca attcttactg
     6361 gaagaaatgc ttagatatgt cgattgaatc cgttatgctt cctcctcctt ggcggttgat
     6421 gccaataact gcagcggctg aagcgatttg ggcgtgttcg attgaagaac tagggctgga
     6481 gaatgaggcc gagaaaccga gcaatgcgtt gttaactaga gcttggtctc caggatggag
     6541 caatgctgat aagttactaa atgagttcat cgagaagcag ttgatagatt atgcaaagaa
     6601 cagcaagaaa gttgttggga attctacttc actactttct ccgtatctcc atttcgggga
     6661 aataagcgtc agacacgttt tccagtgtgc ccggatgaaa caaattatat gggcaagaga
     6721 taagaacagt gaaggagaag aaagtgcaga tctttttctt aggggaatcg gtttaagaga
     6781 gtattctcgg tatatatgtt tcaacttccc gtttactcac gagcaatcgt tgttgagtca
     6841 tcttcggttt ttcccttggg atgctgatgt tgataagttc aaggcctgga gacaaggcag
     6901 gaccggttat ccgttggtgg atgccggaat gagagagctt tgggctaccg gatggatgca
     6961 taacagaata agagtgattg tttcaagctt tgctgtgaag tttcttctcc ttccatggaa
     7021 atggggaatg aagtatttct gggatacact tttggatgct gatttggaat gtgacatcct
     7081 tggctggcag tatatctctg ggagtatccc cgatggccac gagcttgatc gcttggacaa
     7141 tcccgcgtta caaggcgcca aatatgaccc agaaggtgag tacataaggc aatggcttcc
     7201 cgagcttgcg agattgccaa ctgaatggat ccatcatcca tgggacgctc ctttaaccgt
     7261 actcaaagct tctggtgtgg aactcggaac aaactatgcg aaacccattg tagacatcga
     7321 cacagctcgt gagctactag ctaaagctat ttcaagaacc cgtggagcac agatcatgat
     7381 cggagcagca gcccgggatc caccggtcgc caccggagga ggaagcaaag gggaggagtt
     7441 gtgtggggta atgacaatct cggactctcc aagagaacag gatagtgaat cacagacttt
     7501 ggacaaagtt taccaaatga aaagcaaacc tcggggatac tgtctgatca tcaacaatca
     7561 caattttgca aaagcacggg agaaagtgcc caaacttcac agcattaggg acaggaatgg
     7621 aacacacttg gatgcagggg ctttgaccac gacctttgaa gagcttcatt ttgagatcaa
     7681 gccccacgat gactgcacag tagagcaaat ctatgagatt ttgaaaatct accaactcat
     7741 ggaccacagt aacatggact gcttcatctg ctgtatcctc tcccatggag acaagggcat
     7801 catctatggc actgatggac aggaggcccc catctatgag ctgacatctc agttcactgg
     7861 tttgaagtgc ccttcccttg ctggaaaacc caaagtgttt tttattcagg cttgtcaggg
     7921 ggataactac cagaaaggta tacctgttga gactgattca gaggagcaac cctatttaga
     7981 aatggattta tcatcacctc aaacgagata tatcccggat gaggctgact ttctgctggg
     8041 gatggccact gtgaataact gtgtttccta ccgaaaccct gcagagggaa cctggtacat
     8101 ccagtcactt tgccagagcc tgagagagcg atgtcctcga ggcgatgata ttctcaccat
     8161 cctgactgaa gtgaactatg aagtaagcaa caaggatgac aagaaaaaca tggggaaaca
     8221 gatgcctcag cctactttca cactaagaaa aaaacttgtc ttcccttctg attgaaattc
     8281 ccgtatacac cggtgccggc ggatccagac cacctcccct gcgagctaag ctggacagcc
     8341 aatgacgggt aagagagtga catttttcac taacctaaga caggagggcc gtcagagcta
     8401 ctgcctaatc caaagacggg taaaagtgat aaaaatgtat cactccaacc taagacaggc
     8461 gcagcttccg agggatttga gatccagaca tgataagata cattgatgag tttggacaaa
     8521 ccaaaactag aatgcagtga aaaaaatgcc ttatttgtga aatttgtgat gctattgcct
     8581 tatttgtaac cattataagc tgcaataaac aagttaagtc gataaactgg atctctgctg
     8641 tccctgtaat aaacccgaaa attttgaatt tttgtaattt gtttttgtaa ttctttagtt
     8701 tgtatgtctg ttgctattat gtctactatt ctttcccctg cactgtaccc cccaatcccc
     8761 ccttttcttt taaaggcgat accgtcgaga tccgttcact aatcgaatgg atctgtctct
     8821 gtctctctct ccaccttctt cttctattcc ttcgggcctg tcgggtcccc tcggggttgg
     8881 gaggtgggtc tgaaacgata atggtgaata tccctgccta actctattca ctatagaaag
     8941 tacagcaaaa actattctta aacctaccaa gcctcctact atcattatga ataattttat
     9001 ataccacagc caatttgtta tgttaaacca attccacaaa cttgcccatt tatctaattc
     9061 caataattct tgttcattct tttcttgctg gttttgcgat tcttcaatta aggagtgtat
     9121 taagcttgtg taattgttaa tttctctgtc ccactccatc caggtcgtgt gattccaaat
     9181 ctgttccaga gatttattac tccaactagc attccaaggc acagcagtgg tgcaaatgag
     9241 ttttccagag caaccccaaa tccccaggag ctgttgatcc tttaggtatc tttccacagc
     9301 caggattctt gcctggagct gcttgatgcc ccagactgtg agttgcaaca gatgctgttg
     9361 cgcctcaata gccctcagca aattgttctg ctgctgcact ataccagaca ataattgtct
     9421 ggcctgtacc gtcagcgtca ttgacgctgc gcccatagtg cttcctgctg ctcccaagaa
     9481 cccaaggaac aaagctccta ttcccactgc tcttttttct ctctgcacca ctcttctctt
     9541 tgccttggtg ggtgctactc ctaatggttc aatttttact actttatatt tatataattc
     9601 acttctccaa ttgtccctca tatctcctcc tccaggtctg aagatcagcg gccggccgct
     9661 tgctgtgcgg tggtcttact tttgttttgc tcttcctcta tcttgtctaa agcttccttg
     9721 gtgtctttta tctctatcct ttgatgcaca caatagaggg ttgctactgt attatataat
     9781 gatctaagtt cttctgatcc tgtctgaagg gatggttgta gctgtcccag tatttgtcta
     9841 cagccttctg atgtttctaa caggccagga ttaactgcga atcgttctag ctccctgctt
     9901 gcccatacta tatgttttaa tttatatttt ttctttcccc ctggccttaa ccgaattttt
     9961 tcccatcgcg atctaattct cccccgctta atactgacgc tctcgcaccc atctctctcc
    10021 ttctagcctc cgctagtcaa aatttttggc gtactcacca gtcgccgccc ctcgcctctt
    10081 gccgtgcgcg cttcagcaag ccgagtcctg cgtcgagaga gctcctctgg tttccctttc
    10141 gctttcaagt ccctgttcgg gcgccactgc tagagatttt ccacactgac taaaagggtc
    10201 tgagggatct ctagttacca gagtcacaca acagacgggc acacactact tgaagcactc
    10261 aaggcaagct ttattgaggc ttaagcagtg ggttccctag ttagccagag agctcccagg
    10321 ctcagatctg gtctaaccag agagacccag tacaggcaaa aagcagctgc ttatatgcag
    10381 gatctgaggg ctcgccactc cccagtcccg cccaggccac gcctccctgg aaagtcccca
    10441 gcggaaagtc ccttgtagca agctcgatat cagcagttct tgaagtactc cggatgcagc
    10501 tctcgggcca cgtgatgaaa tgctaggcgg ctgtcaaacc tccactctaa cacttctctc
    10561 tccgggtcat ccatcccatg caggctcaca gggtgtaaca agctggtgtt ctctccttta
    10621 ttggcctctt ctaccttatc tggctcaact ggtactagct tgtagcacca tccaaaggtc
    10681 agtggatatc tgacccctgg ccctggtgtg tagttctgct aatcagggaa gtagccttgt
    10741 gtgtggtaga tccacagatc aaggatatct tgtcttcttt gggagtgaat tagcccttcc
    10801 aactactaag tttgtagtac atatttaaca aatacaattt ctttaaaatg aaaataattc
    10861 agaggaatca caggtttaga gtaaatgaaa ccacaggtaa ttggcagtgg taatagggta
    10921 tggggtggga agtttgggat gattttggtt agcttgagtt atccagttga tccagacatg
    10981 ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt
    11041 atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa
    11101 gttgatctcc cgatccgtcg acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc
    11161 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
    11221 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
    11281 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
    11341 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
    11401 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
    11461 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
    11521 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
    11581 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
    11641 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
    11701 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
    11761 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
    11821 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
    11881 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
    11941 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
    12001 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
    12061 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
    12121 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
    12181 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
    12241 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttc
//
