LOCUS       sequence-460772-       10275 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Tet inducible expression of DmrB-Mlkl with mCherry to induce 
            phosphomimetic MLKL S345D necroptosis.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10275)
  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 10275)
  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: pLVX MLKL(S345D)-2A-mCherry-puro
FEATURES             Location/Qualifiers
     source          1..10275
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(161..205)
                     /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    390..623
                     /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    1120..1245
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             1292..1925
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     polyA_signal    2093..2227
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2117..2136)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2171..2190
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        2275..2379
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2380..2448,2449..3240)
                     /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(2598..2617)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3411..3999
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3900..3919
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4153..4170
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4287..4308
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4323..4328,4329..4346,4347..4353)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4361..4377
                     /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     4366..4388
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4385..4401
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4385..4401
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     LTR             4529..5162
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    5369..5957
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             5475..5486
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     5884..5904
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5974..6573)
                     /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(6057..6077)
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     6554..6573
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(6594..7093)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(6642..6661)
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     primer_bind     7035..7058
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     CDS             complement(7127..7447)
                     /codon_start=1
                     /product="F36V mutant of FK506-binding protein FKBP12"
                     /label=DmrB
                     /note="binds synthetic ligands that do not bind wild-type 
                     FKBP"
                     /translation="GVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKVDSSRDRNKP
                     FKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELL
                     KLE"
     CDS             complement(7471..8181)
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(7749..7768)
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     8018..8036
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     CDS             complement(8182..8235)
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     promoter        complement(9646..9995)
                     /label=TRE3GV promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G, optimized for 
                     retroviral and lentiviral vectors"
     primer_bind     complement(9683..9707)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    9753..9771
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    9789..9807
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    9825..9843
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    9861..9879
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    9897..9915
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    9933..9951
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    9969..9987
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     misc_feature    10058..10173
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1 (lacking the first T)"
ORIGIN
        1 ttgggaggtg ggtctgaaac gataatggtg aatatccctg cctaactcta ttcactatag
       61 aaagtacagc aaaaactatt cttaaaccta ccaagcctcc tactatcatt atgaataatt
      121 ttatatacca cagccaattt gttatgttaa accaattcca caaacttgcc catttatcta
      181 attccaataa ttcttgttca ttcttttctt gctggttttg cgattcttca attaaggagt
      241 gtattaagct tgtgtaattg ttaatttctc tgtcccactc catccaggtc gtgtgattcc
      301 aaatctgttc cagagattta ttactccaac tagcattcca aggcacagca gtggtgcaaa
      361 tgagttttcc agagcaaccc caaatcccca ggagctgttg atcctttagg tatctttcca
      421 cagccaggat tcttgcctgg agctgcttga tgccccagac tgtgagttgc aacagatgct
      481 gttgcgcctc aatagccctc agcaaattgt tctgctgctg cactatacca gacaataatt
      541 gtctggcctg taccgtcagc gtcattgacg ctgcgcccat agtgcttcct gctgctccca
      601 agaacccaag gaacaaagct cctattccca ctgctctttt ttctctctgc accactcttc
      661 tctttgcctt ggtgggtgct actcctaatg gttcaatttt tactacttta tatttatata
      721 attcacttct ccaattgtcc ctcatatctc ctcctccagg tctgaagatc agcggccggc
      781 cgcttgctgt gcggtggtct tacttttgtt ttgctcttcc tctatcttgt ctaaagcttc
      841 cttggtgtct tttatctcta tcctttgatg cacacaatag agggttgcta ctgtattata
      901 taatgatcta agttcttctg atcctgtctg aagggatggt tgtagctgtc ccagtatttg
      961 tctacagcct tctgatgttt ctaacaggcc aggattaact gcgaatcgtt ctagctccct
     1021 gcttgcccat actatatgtt ttaatttata ttttttcttt ccccctggcc ttaaccgaat
     1081 tttttcccat cgcgatctaa ttctcccccg cttaatactg acgctctcgc acccatctct
     1141 ctccttctag cctccgctag tcaaaatttt tggcgtactc accagtcgcc gcccctcgcc
     1201 tcttgccgtg cgcgcttcag caagccgagt cctgcgtcga gagagctcct ctggtttccc
     1261 tttcgctttc aagtccctgt tcgggcgcca ctgctagaga ttttccacac tgactaaaag
     1321 ggtctgaggg atctctagtt accagagtca cacaacagac gggcacacac tacttgaagc
     1381 actcaaggca agctttattg aggcttaagc agtgggttcc ctagttagcc agagagctcc
     1441 caggctcaga tctggtctaa ccagagagac ccagtacagg caaaaagcag ctgcttatat
     1501 gcaggatctg agggctcgcc actccccagt cccgcccagg ccacgcctcc ctggaaagtc
     1561 cccagcggaa agtcccttgt agcaagctcg atatcagcag ttcttgaagt actccggatg
     1621 cagctctcgg gccacgtgat gaaatgctag gcggctgtca aacctccact ctaacacttc
     1681 tctctccggg tcatccatcc catgcaggct cacagggtgt aacaagctgg tgttctctcc
     1741 tttattggcc tcttctacct tatctggctc aactggtact agcttgtagc accatccaaa
     1801 ggtcagtgga tatctgaccc ctggccctgg tgtgtagttc tgctaatcag ggaagtagcc
     1861 ttgtgtgtgg tagatccaca gatcaaggat atcttgtctt ctttgggagt gaattagccc
     1921 ttccaactac taagtttgta gtacatattt aacaaataca atttctttaa aatgaaaata
     1981 attcagagga atcacaggtt tagagtaaat gaaaccacag gtaattggca gtggtaatag
     2041 ggtatggggt gggaagtttg ggatgatttt ggttagcttg agttatccag ttgatccaga
     2101 catgataaga tacattgatg agtttggaca aaccacaact agaatgcagt gaaaaaaatg
     2161 ctttatttgt gaaatttgtg atgctattgc tttatttgta accattataa gctgcaataa
     2221 acaagttgat ctcccgatcc gtcgacgtca ggtggcactt ttcggggaaa tgtgcgcgga
     2281 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     2341 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     2401 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     2461 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     2521 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     2581 agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag
     2641 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     2701 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     2761 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     2821 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     2881 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg
     2941 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     3001 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     3061 tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg
     3121 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     3181 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     3241 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     3301 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     3361 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     3421 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     3481 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     3541 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     3601 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     3661 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     3721 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     3781 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     3841 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     3901 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     3961 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     4021 ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc gttatcccct
     4081 gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg ccgcagccga
     4141 acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat acgcaaaccg
     4201 cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt tcccgactgg
     4261 aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta ggcaccccag
     4321 gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt
     4381 cacacaggaa acagctatga ccatgattac gccaagctct agctagaggt cgacggtaaa
     4441 cgctagcaat gtcaaggcct ctcactctct gatattcatt tctttgcaag ttataaatac
     4501 tgaataataa gatgacatga actactactg ctagagattt tccacactga ctaaaagggt
     4561 ctgagggatc tctagttacc agagtcacac aacagacggg cacacactac ttgaagcact
     4621 caaggcaagc tttattgagg cttaagcagt gggttcccta gttagccaga gagctcccag
     4681 gctcagatct ggtctaacca gagagaccca gtacaggcaa aaagcagctg cttatatgca
     4741 ggatctgagg gctcgccact ccccagtccc gcccaggcca cgcctccctg gaaagtcccc
     4801 agcggaaagt cccttgtagc aagctcgata tcagcagttc ttgaagtact ccggatgcag
     4861 ctctcgggcc acgtgatgaa atgctaggcg gctgtcaaac ctccactcta acacttctct
     4921 ctccgggtca tccatcccat gcaggctcac agggtgtaac aagctggtgt tctctccttt
     4981 attggcctct tctaccttat ctggctcaac tggtactagc ttgtagcacc atccaaaggt
     5041 cagtggatat ctgacccctg gccctggtgt gtagttctgc taatcaggga agtagccttg
     5101 tgtgtggtag atccacagat caaggatatc ttgtcttcgt tgggagtgaa ttagcccttc
     5161 cagtcccctc ttttctttta aaaagtggct aagatctaca gctgccttgt aagtcattgg
     5221 tcttaaaggt acctgaggtg tgactggaaa acccacctcc tcctcctctt gtgcttctag
     5281 ccaggcacaa tcagcattgg tagctgctgt attgctactt gtgattgctc catgtttttc
     5341 taggtctcga ctgcagaatt aattccaggc ggggaggcgg cccaaaggga gatccgactc
     5401 gtctgagggc gaaggcgaag acgcggaaga ggccgcagag ccggcagcag gccgcgggaa
     5461 ggaaggtccg ctggattgag ggccgaaggg acgtagcaga aggacgtccc gcgcagaatc
     5521 caggtggcaa cacaggcgag cagccatgga aaggacgtca gcttccccga caacaccacg
     5581 gaattgtcag tgcccaacag ccgagcccct gtccagcagc gggcaaggca ggcggcgatg
     5641 agttccgccg tggcaatagg gagggggaaa gcgaaagtcc cggaaaggag ctgacaggtg
     5701 gtggcaatgc cccaaccagt gggggttgcg tcagcaaaca cagtgcacac cacgccacgt
     5761 tgcctgacaa cgggccacaa ctcctcataa agagacagca accaggattt atacaaggag
     5821 gagaaaatga aagccatacg ggaagcaata gcatgataca aaggcattaa agcagcgtat
     5881 ccacatagcg taaaaggagc aacatagtta agaataccag tcaatctttc acaaattttg
     5941 taatccagag gttgattgtt ccagacgcgc ccgtcaggca ccgggcttgc gggtcatgca
     6001 ccaggtgcgc ggtccttcgg gcacctcgac gtcggcggtg acggtgaagc cgagccgctc
     6061 gtagaagggg aggttgcggg gcgcggaggt ctccaggaag gcgggcaccc cggcgcgctc
     6121 ggccgcctcc actccgggga gcacgacggc gctgcccaga cccttgccct ggtggtcggg
     6181 cgagacgccg acggtggcca ggaaccacgc gggctccttg ggccggtgcg gcgccaggag
     6241 gccttccatc tgttgctgcg cggccagccg ggaaccgctc aactcggcca tgcgcgggcc
     6301 gatctcggcg aacaccgccc ccgcttcgac gctctccggc gtggtccaga ccgccaccgc
     6361 ggcgccgtcg tccgcgaccc acaccttgcc gatgtcgagc ccgacgcgcg tgaggaagag
     6421 ttcttgcagc tcggtgaccc gctcgatgtg gcggtccgga tcgacggtgt ggcgcgtggc
     6481 ggggtagtcg gcgaacgcgg cggcgagggt gcgtacggcc ctggggacgt cgtcgcgggt
     6541 ggcgaggcgc accgtgggct tgtactcggt catggtaagc ttgggctgca ggtcgaaagg
     6601 cccggagatg aggaagagga gaacagcgcg gcagacgtgc gcttttgaag cgtgcagaat
     6661 gccgggcctc cggaggacct tcgggcgccc gccccgcccc tgagcccgcc cctgagcccg
     6721 cccccggacc caccccttcc cagcctctga gcccagaaag cgaaggagca aagctgctat
     6781 tggccgctgc cccaaaggcc tacccgcttc cattgctcag cggtgctgtc catctgcacg
     6841 agactagtga gacgtgctac ttccatttgt cacgtcctgc acgacgcgag ctgcggggcg
     6901 ggggggaact tcctgactag gggaggagta gaaggtggcg cgaaggggcc accaaagaac
     6961 ggagccggtt ggcgcctacc ggtggatgtg gaatgtgtgc gaggccagag gccacttgtg
     7021 tagcgccaag tgcccagcgg ggctgctaaa gcgcatgctc cagactgcct tgggaaaagc
     7081 gcctccccta cccggtatca gtcagtcaga attccatatg acgcgtttcc agttttagaa
     7141 gctccacatc gaagacgaga gtggcatgtg gtgggatgat gcctgggtgc ccagtggcac
     7201 cataggcata atctggagat atagtcagtt tggctctctg acccacactc atctgggcaa
     7261 ccccttcttc ccagcctcgg atcacctcct gcttgcctag cataaactta aagggcttgt
     7321 ttctgtcccg ggaggaatca actttctttc catcttcaag catcccggtg tagtgcacca
     7381 cgcaggtctg gccgcgcttg gggaaggtgc gcccgtctcc tggggagata gtctccacct
     7441 gcactcctct agaagccatg cggccgcggg tcacttgtac agctcgtcca tgccgccggt
     7501 ggagtggcgg ccctcggcgc gttcgtactg ttccacgatg gtgtagtcct cgttgtggga
     7561 ggtgatgtcc aacttgatgt tgacgttgta ggcgccgggc agctgcacgg gcttcttggc
     7621 cttgtaggtg gtcttgacct cagcgtcgta gtggccgccg tccttcagct tcagcctctg
     7681 cttgatctcg cccttcaggg cgccgtcctc ggggtacatc cgctcggagg aggcctccca
     7741 gcccatggtc ttcttctgca ttacggggcc gtcggagggg aagttggtgc cgcgcagctt
     7801 caccttgtag atgaactcgc cgtcctgcag ggaggagtcc tgggtcacgg tcaccacgcc
     7861 gccgtcctcg aagttcatca cgcgctccca cttgaagccc tcggggaagg acagcttcaa
     7921 gtagtcgggg atgtcggcgg ggtgcttcac gtaggccttg gagccgtaca tgaactgagg
     7981 ggacaggatg tcccaggcga agggcagggg gccacccttg gtcaccttca gcttggcggt
     8041 ctgggtgccc tcgtaggggc ggccctcgcc ctcgccctcg atctcgaact cgtggccgtt
     8101 cacggagccc tccatgtgca ccttgaagcg catgaactcc ttgatgatgg ccatgttatc
     8161 ctcctcgccc ttgctcacca tagggccggg attctcctcc acgtcaccgc atgttagaag
     8221 acttcctctg ccctcgggcc ccaccttctt gtccgtggat tcttcaaccg cagacagtct
     8281 ctccaagatt ccgtccacag agggccgttg ggagggctca tgggcacgac actcattaat
     8341 gatttcccgc aacaactcag ggcaatcctg acccactggt tcctgcttct tgtcctcagc
     8401 caccagctcc cggatcttct tagaatcaca gccttcaaat gggatctttc cagtggcaat
     8461 ttcccagagt acaattccaa agctatatat ttcagctttt atgtcataaa ggcaaaatgg
     8521 atttttcagt ctctcagggg agacatatat cgttgaactg gatctctctg ctttagtgct
     8581 ctttgctgtc cggctgatgt cattctgtgt tttgcttaac tcaaatcctg caagctttac
     8641 ttggtagcct ccggctacga ggaaactgga gctgctgatg tttctgtgga gtgtttccga
     8701 atggtgtagc ctgtataagc ctctggctgc cctcaggact aggaggctgc gcacactcat
     8761 tgtgaggtct ttttctctat ccagcagttc cctcagggtt ccaagttcac agtactccat
     8821 gacaatggag aactcagggg gcttcactgt ttgatcaatg caaatcccaa atatacgcaa
     8881 gatgttggga gaatcgaatt tcttcatggt tttgatctcg tcattgaaag tgaacctcac
     8941 tattccaaca ctttcggcct gggggttgtt gaatactttg atggtaactg gagatctgtg
     9001 atactctcct ctataaatgg tgctcatttt acttgtcttc agtttggtcc acggaggtcc
     9061 aagatgttcc tttggaatct ccttgatttg gagatcttgt gggatctcct gtgtgggttt
     9121 tagtgagcat tgcttcaggg ttttgttgat ttcttccacg ctaatttgca actgcatcag
     9181 gataactttc atattttcat ttccgtcttc ctctgcatcc tgtcgatctt cctgctgcca
     9241 ggatgctggc tggctgacat ctgaaacggt attccaatga taaacctgaa gcagcagcaa
     9301 cagctcctcc cagacgtctc tcagcttctc attcacttca tggaagagga tcttatcatt
     9361 gcccacactc acaaacttcc aaatatggga cttcttgctg aacttttcta tctgctggtt
     9421 agcctccttc aggacttcat caaaacggcc cagggcagca gtaatgtcat cgggcaggtt
     9481 cttctttcct tgggcctgga gtctctggag aggctgtagc aggccgtgca cacggtttcc
     9541 tagacgctgg cattgtttcc ggcagtattt catcttttca cactgttcat agatgagctg
     9601 gcctaacttg atgatctgtc ccaatttatc catgggcccg gatccaagta taagacaaaa
     9661 gtgttgtgga attgctccag gcgatctgac ggttcactaa acgagctctg cttttatagg
     9721 cgcccaccgt acacgcctac ctcgacatac gttctctatc actgataggg agtaaactgg
     9781 atatacgttc tctatcactg atagggagta aactgtagat acgttctcta tcactgatag
     9841 ggagtaaact ggtcatacgt tctctatcac tgatagggag taaactcctt atacgttctc
     9901 tatcactgat agggagtaaa gtctgcatac gttctctatc actgataggg agtaaactct
     9961 tcatacgttc tctatcactg atagggagta aactcgagtg aagacgaaag ggcctcatcg
    10021 ataaactgga tctctgctgt ccctgtaata aacccgaaaa ttttgaattt ttgtaatttg
    10081 tttttgtaat tctttagttt gtatgtctgt tgctattatg tctactattc tttcccctgc
    10141 actgtacccc ccaatccccc cttttctttt aaaggcgata ccgtcgagat ccgttcacta
    10201 atcgaatgga tctgtctctg tctctctctc caccttcttc ttctattcct tcgggcctgt
    10261 cgggtcccct cgggg
//
