| General information | |||
| TC004749 | Gene
The sequence from the iB fragment is matched against the genes from the official gene set. The matche(s) is/are visualizied in a genome browser. |
NC_007416.3: 6357346..6370599 | |
| Sequence information | |||
| Transcripts / Proteins (1) | Get mRNA sequence
Get CDS sequence Get protein sequence |
||
| Ortholog information (from OrthoDB)
The information about the homolog genes in Drosophila
is provided by OrthoDB http://www.orthodb.org/.
The homologs are sorted according to their similarity score. For an orthology statement additional
analysis are required. | |||
| Closest fly homologs |
|
TC004749 | |
Attention! iBeetle is a first pass screen. Hence, the documented phenotypes need to be confirmed by independent experiments using non-overlapping dsRNA fragments! Phenotypes with a penetrance > 50% are frequently reproduced. See Schmitt-Engel et al. 2015 for details on reproducibility.
| General information | ||||
| iB_03833 | iB fragment
The sequence from the iB fragment is matched against the genes from the official gene set. The matche(s) is/are visualizied in a genome browser. |
Get FastA Get primers |
||
Usually 10 injected animals
Pupal injection procedure
Day 0: 10 female pupae of the pBA19 strain (muscle enhancer trap line) were injected with dsRNA.
3 days post injection (dpi): Hatch control: Pupal and adult lethality as well as metamorphosis defects (molting, eclosion) were documented. For mating, 4 males of the black strain were added.
9 dpi: First egg-lay was collected and incubated for cuticle analysis. Adult lethality and egg production (reduced/ no egg-lay) was documented.
11 dpi: Second egg-lay was collected and incubated for embryonic muscle analysis. Adult lethality and egg production (reduced/ no egg-lay) were documented.
Note: The adult morphology was not analyzed systematically at 3/9/11 dpi. Only obvious phenotypes, visible without magnification, have been annotated.
13 dpi: Egg productivity and Ovary analysis
The percentage of hatched larvae was documented and not hatched larvae/ eggs were embedded for cuticle analysis (15 dpi).
In case of a reduction of egg production, 4 injected females were dissected to analyze the gross morphology of the ovaries.
14 dpi: Analysis of embryonic musculature and early embryonic development
Offspring of the injected females (hatched and not hatched larvae/ eggs) were analysed for embryonic lethality and muscle defects.
15 dpi: Analysis of larval instar 1 cuticle
Offspring of injected females were analysed and cuticle phenotypes were annotated.
22 dpi: Stink gland analysis
Documentation of defects in abdominal and thoracic stink glands (colour, size, content) of the injected femals.
Terms used in the pupal injection procedure
- larval pantagmatic defects
- At least two tagmata (head, thorax, abdomen or terminus) show similar/ comparable defects.
- empty eggs
- Just empty shells are visible, no cuticle has been developed.
- strong defects
- L1 cuticle larvae which show severe defects. These strong defects are subdivided into three categories
- strongly fragmented (cuticle crumbs)
- No segments or appendages are identifiable, only residues of cuticle and bristles can be identified.
- cuticle remnants (with unidentifiable segments)
- Segments or parts of appendages are recognizable, but identity is not clear.
- cuticle remnants (with some identifiable segments)
- At least one segment or appendage is clearly regonizable.
- number of eggs on slide
- affected embryos usually (not always) do not hatch. Only the not hatched were used for cuticle analysis.
For the embryonic muscle analysis the hatched as well as the not hatched were analysed. - total number of affected eggs/embryos/larvae
- Summary of all different phenotypes/ phenotypic series which were annotated.
- inside-out
- The L1 larva shows a partial or complete inversion., e.g. bristles, appendages or parts of the abdominal segments are inverted into the interior of the cuticle.
- starved
- Dissected females show a strong resorption of fatbody predominately in the abdomen and the oogenesis is blocked. These are considered as eventually lethal phenotypes and the blocked oogenesis probably a secondary defect due to starvation.
- eclosion not fulfilled
- The emergence of the adult from the pupa stage is interrupted. This phenotype shows pupal as well as adult features.
Metamorphosis and survival
Lethality 11 days after pupal injection: 20.0% (includes death as pupa, adult)
eclosion not fulfilled (with pupal and adult features) - Number of animals showing the phenotype: 1 (3 dpi)Days post injection
adult lethality - Number of animals showing the phenotype: 1 (9 dpi)
Days post injection
Analysis of embryonic musculature and early development
number of eggs/embryos/larvae on slide: > 50
embryo/egg with muscle fluorescence/pattern, but not hatched (pot. late embryonic lethality) -
Penetrance (percentage of animals/eggs on slide): 50% - 80%
larval musculature irregular - Penetrance (percentage of animals/eggs on slide): less than 30%
affected eggs/embryos/larvae total percentage of affected eggs/embryos/larvae on slide - Penetrance (percentage of animals/eggs on slide): 50% - 80%
remarks: muscle pattern potentially obscured by segmentation defect,
larval musculature irregular - Penetrance (percentage of animals/eggs on slide): less than 30%
affected eggs/embryos/larvae total percentage of affected eggs/embryos/larvae on slide - Penetrance (percentage of animals/eggs on slide): 50% - 80%
remarks: muscle pattern potentially obscured by segmentation defect,
Analysis of larval stage 1 cuticle
number of eggs/embryos/larvae on slide: > 50
larva random partiallyconstricted - Penetrance (percentage of animals/eggs on slide): less than 30%
abdomen partially circumference constricted - Penetrance (percentage of animals/eggs on slide): less than 30%
abdomen partially constricted - Penetrance (percentage of animals/eggs on slide): less than 30%
affected eggs/embryos/larvae total percentage of affected eggs/embryos/larvae on slide - Penetrance (percentage of animals/eggs on slide): less than 30%
larva random partiallyconstricted - Penetrance (percentage of animals/eggs on slide): less than 30%
abdomen partially circumference constricted - Penetrance (percentage of animals/eggs on slide): less than 30%
abdomen partially constricted - Penetrance (percentage of animals/eggs on slide): less than 30%
affected eggs/embryos/larvae total percentage of affected eggs/embryos/larvae on slide - Penetrance (percentage of animals/eggs on slide): less than 30%
Usually 10 injected animals
Larval injection procedure
Day 0: 10 female L5-6 larvae of the D17Xhom strain (females express RFP in the eyes and in the CNS; pupae express GFP in the thorax) were injected with dsRNA.
11 days post injection (dpi): Pupal morphology Documentation of larval/ pupal death and defects of metamorphosis (eclosion, delayed development). Screening for alterations of pupal head and appendages, pupal thorax, pupal thoracic musculature, pupal legs, pupal wings, pupal abdomen and pupal genital lobes.
16 dpi: Adult morphology Documentation of larval/ pupal and adult death and defects of metamorphosis (eclosion, delayed development). Screening for alterations of adult head and appendages, adult thorax, adult elytra and hindwings, adult legs and cuticle properties. For mating 4 males of the black strain were added.
19 dpi: Sieving and egg-lay Documentation of adult lethality and enduring metamorphosis defects.
22 dpi: Ovary analysis (Fertility) In case of a reduction of egg production 4 females were dissected and the gross morphology of the ovaries were analysed.
41 dpi: Fertility and stink gland analysis Documentation of defects in abdominal and thoracic stink glands development (colour, size, content) of the injected females. Check for fertility of injected larvae (no or decreased offspring).
Terms used in the larval injection procedure
- pupal/ adult pantagmatic defects
- At least two tagmata (head, thorax, abdomen or terminus) show similar/ comparable defects.
- pupal molt not fulfilled
- The hatching from the larva to the pupa is interrupted. Either it dies as prepupa or the phenotype shows larval as well as pupal features.
- eclosion not fulfilled
- The emergence of the adult from the pupa stage is interrupted. This phenotype shows pupal as well as adult features.
- starved
- Dissected females show a strong resorption of fatbody predominately in the abdomen and the oogenesis is blocked. These are considered as eventually lethal phenotypes and the blocked oogenesis probably a secondary defect due to starvation.
Metamorphosis and survival
Lethalities 11 days after larval injection: 10.0% (includes death as larva, prepupa, pupa)
Lethalities 22 days after larval injection: 10.0% (includes death as larva, prepupa, pupa, adult)
Days post injection
adult lethality - Number of animals showing the phenotype: 2 (41 dpi)
Days post injection
Fertility
egg production: number no/almost no eggs -
mature egg deposition >5 number 2nd lateral oviduct: number 1st lateral oviduct: blocked - Number of animals showing the phenotype: 4
mature egg deposition >5 number 2nd lateral oviduct: number 1st lateral oviduct: blocked - Number of animals showing the phenotype: 4
- >TC004749 RA
MSVASDYSS EEELSLFRP FTRESLAVI ETRIAEEYA KQKELEKKR AEGEIRYED EDEDEGPQP DPTLEQGLP LPVRLHSTF PPELASTPL EDIDPFYSN QRTFVVVSK GKDIFRFSA TNALWVLDP FNPIRRVAI YILVHPLFS LFIITTILV NCILMIMPT TPTVESTEV IFTGIYTFE SAVKVMARG FILQPFTYL RDAWNWLDF VVIALAYVT MGIDLGNLA ALRTFRVLR ALKTVAIVP GLKTIVGAV IESVKNLRD VIILTMFSL SVFALIGLQ IYMGVLTQK CIKEFPLDG SWGNLTHEN WERFVSNES NWYWDDEKG ERPLCGNSS GSGMCKEGY VCLQGYGDN PNYGYTSFD TFGWALLSA FRLMTQDYW ENLYQLVLR SAGPWHMLF FIVIIFLGS FYLVNLILA IVAMSYDEL QKKAQDEEE AEANAIREA EQKAKDKQD RADARAAAA VEAAEAAAA AAAAAAAGC DNDQYIIKS SSDYSCHSY ESFVGQPNG QDDNNKEKI SIRSEGLDS VSEQRRVPT NLSKMRKVS AASLSLPGS PFNLRRGSR GSHQFTLRN NRRMVAPPG DRKPLVLST YLDAQEHLP YADDSNAVT PMSEENGVM VLPMYYTNL GSRHSSYTS HASRMSYTS HGDLLGGLG SNGKMMTKE SQLRNRSLR NGPAAAPLN HKSTRGVYD GPTGHICES KVKQIDNPF IDSNQRQTV VDMKDVMVL NDIIEQAAV SGGSDRGAN NDDEEEEPT FRERFLAYA IKIIDMFCV WDCCKCWVV IQKFVALLV FDPFVELFI TLCIVVNTL FMALDHHDM DKDLEKALK SGNYFFTAT FMIEATLKL IAMSPKYYF QEGWNIFDF IIVALSLLE LGLEGVQGL SVLRSFRLL RVFKLAKSW PTLNLLISI MGRTMGALG NLTFVLCII IFIFAVMGM QLFGKNYTD NVDRFPDHE LPRWNFTDF MHSFMIVFR VLCGEWIES MWDCMLVGD VSCIPFFLA TVVIGNLVV LNLFLALLL SNFGSSSLS APTADNDTN KIAEAFDRI GRFIRWIKG NVLHIAKLV RFKLTNQIS DQPSDVRCD GGLDISGDE ILADGIIKK SPKDRLEVT IGTGMDLTV HGDPKRGKN NINKSKTIG NSILEHGMY IGHLDEDEI SNKSYGSHK NRFKEESHK GSVDMLDDH EEKRDASKE ELGIGEEET EDDRFDLEK PLTDIVIVG NADDIIIDD YAADCFPDS CYKKFPFLA GDEDSPFWQ GWGNLRSKT YRLIENKYF ETAVITMIL LSSMALALE DVHLQSRPI LQDILYYMD RIFTVIFFF EMLIKWLAL GFQKYFTNA WCWLDFVIV MVSLINFIA SLCGAGGIQ AFKTMRTLR ALRPLRAMS RMQGMRVVV NALVQAIPS IFNVLLVCL IFWLIFAIM GVQLFAGKY FKCVDANKT TLSYEIIPD VNACKAENY TWENSPMNF DHVGKAYLC LFQVATFKG WIQIMNDAI DSRDINKQP IRETNIYMY LYFVFFIIF GSFFTLNLF IGVIIDNFN EQKKKGGDS LALMMTDAQ KKYYNAMKK MGSKKPMKA IPRPRWRPQ AIVFEIVTN KKFDMIIML FIGLNMLTM TMDHYQQKE TFTKVLDSL NMIFIVIFS TECLMKIFA LRYHYFTEP WNLFDLVVV ILSILGLVL SDIIEKYFV SPTLLRVVR VAKVGRVLR LVKGAKGIR TLLFALAMS LPALFNICL LLFLVMFIF AIFGMSFFM HVKDKSGLD DVYNFKTFG QSMILLFQM STSAGWDGV LDGIINEED CKPPNSEIG ETGNCGNST IGIAFLLSY LVISFLIVI NMYIAVILE NYSQATEDV QEGLTDDDY DMYYEIWQQ FDPDGTQYI RYDQLSDLF DVLEPPLQI HKPNKFKIY SMDIPICKG DMMFCVDIL DALTKDFFA RKGNEIEET AELSEVQPR QNEPGYEQV SSTLYRQRE EYCARLIQN AWRKYRQRT GGAPEQVNV DGAAGEGDG EVEVHQTAV LVESDGFVT KNGHKVVLH SRTPSISSK SADV
- >TC004749 RA
atgtccgtt gcctctgac tattcgtcc gaggaggag ctcagccta ttccgtcca ttcactcgc gagtctctt gcggtcatc gagacgaga attgcggaa gaatatgct aagcagaag gagctcgaa aagaaaaga gctgaagga gagattcgg tatgaggac gaggacgaa gatgaaggt ccccagcct gatccaact ctggagcag gggctgccc ttgccagta cgtcttcac agtaccttt cctccggaa ctggccagt acaccactt gaagacatc gaccctttt tacagtaac caaaggact tttgtagtg gttagtaaa ggcaaggac attttccgg tttagcgcc acaaacgcc ctgtgggta ctcgatccc ttcaatccg atccgtcga gtggccatt tacattcta gtgcatcct ttgttttct ctttttatc attaccaca atcctcgtt aactgtatc cttatgatc atgcccaca acacccacc gtagagtca accgaagtg atattcaca ggcatctac acgtttgaa tcagctgtg aaggtgatg gcgagaggt tttatccta caaccgttc acctacctt agagatgca tggaactgg ctcgacttc gtagtcata gctttagct tatgtcacc atgggtata gaccttggg aatcttgct gccttgaga acatttagg gtactacga gctctaaaa actgttgcc attgtgcca ggtttgaag actatcgtg ggagctgta atagaatct gtaaaaaat ctccgagat gtgataatt ttaacaatg ttttccctg tcagtgttc gcattgata ggtttacaa atttatatg ggtgtttta acacaaaaa tgtatcaaa gaatttcct ctggatggc tcttggggg aacctcacc cacgaaaac tgggagaga tttgttagc aacgaatcc aactggtat tgggatgac gaaaaaggc gaaaggcct ctgtgtgga aactcctca ggatcggga atgtgcaaa gagggttac gtttgtttg caaggttac ggtgataac ccaaactat ggttacacc agctttgat acttttggt tgggcatta ctttccgcc ttcagatta atgactcag gattattgg gagaattta taccaactg gttttaaga tcagccggt ccttggcat atgctattt tttatcgtg attatattt ttgggttcg ttttattta gttaacttg atcttggct atcgtcgct atgtcgtac gacgagttg cagaagaaa gcccaagat gaggaagaa gctgaagct aacgctata agggaagcc gagcaaaag gcaaaggac aagcaagat cgggcggat gcgagggca gcggcggcc gtggaggcg gccgaggca gcggccgct gccgcggcg gcggcggca gcagggtgt gacaacgac cagtacatc ataaaatcg tcttcggac tactcgtgt cacagctat gagtcgttc gtggggcaa cccaacggt caagacgac aacaacaag gagaagatt agcatacgc tcggaagga ttagattcc gtgagtgaa cagagaaga gttcctact aatctgagc aagatgcgg aaagtcagc gctgccagt ctcagcctc cccggctct cccttcaac cttcggcga gggtcccga ggtagccac cagttcacc ttgcggaac aataggcgc atggtggct ccgccgggc gatcgaaag cccttggtg ctgtcgacc taccttgac gcccaagag cacctcccc tacgccgac gactctaac gccgtcacc cccatgagc gaggagaac ggcgtcatg gtcctgccc atgtactac accaacctg ggatcacgt cattcatcg tacacttca catgcctca cggatgtca tacacttct catggagat ttgttaggc ggtttggga agcaatggc aaaatgatg acgaaggag agccagctc aggaaccga tcgctgaga aacggacca gcggccgct cccctaaac cacaagtca acgcgaggc gtctacgat ggtcccacg gggcacatt tgcgaatca aaagttaag caaattgac aatcctttt atcgatagc aatcagcga cagacagtc gtggacatg aaagatgtt atggtgctg aacgacatc atcgagcaa gctgccgtg agcggaggc agtgaccgc ggagccaat aatgacgac gaagaggaa gagccgacg ttcagggaa cgcttcctg gcttatgcc atcaaaatc attgacatg ttttgcgta tgggattgt tgtaaatgt tgggtcgtt attcagaag tttgtcgcc cttcttgtc ttcgaccca ttcgtcgag ctttttatc acgctgtgc attgttgtt aacaccttg tttatggca ttggatcac cacgacatg gacaaggat ttggaaaag gctttgaag agcggaaac tactttttt actgcgact tttatgatt gaagctact ttaaaattg attgcaatg agtccgaaa tactacttt caagaaggc tggaacatt tttgatttt attatagtg gcgttatcg ttgctggag ctaggactg gaaggcgtt caaggctta tcggtcttg cgttcattt cgactgtta cgagtgttc aaactggcc aagtcgtgg ccaacttta aatttactc atttccata atgggtaga acgatgggt gctttagga aacttgact tttgtgctc tgtattata atatttatt tttgccgtt atgggcatg caactgttt gggaaaaat tacacagat aacgtggat cggtttccc gaccacgag ctcccccgc tggaacttc accgacttc atgcattcg tttatgata gtttttcgc gttttgtgt ggcgaatgg atcgagtcc atgtgggac tgtatgctg gtgggcgac gtctcttgc attccgttc tttctcgca actgtcgtg attggcaat cttgtggtc cttaatctt ttcttagcc ctgcttttg agtaacttt ggatcttcg agtctgtcg gcgccgact gccgacaat gacacgaat aaaatagca gaggctttt gacagaatc ggccgcttt atcagatgg atcaaaggc aatgtttta catatcgca aaactggtc cgattcaag ctcaccaac caaatttcc gaccagccg tcagatgtg agatgtgat ggtggttta gatatctcc ggagacgaa attttagcc gacggtata attaaaaaa agtcctaaa gaccgactg gaagtaaca attggcacc ggaatggat cttaccgtg catggtgat cccaaaaga ggcaaaaac aatattaac aaaagcaaa acaatcgga aattcaatt ctggaacat ggaatgtac atcggtcac ctggacgaa gacgaaatt agtaacaaa tcgtacggt agtcacaaa aatcgattt aaagaggaa agtcataaa gggagtgtt gatatgtta gacgaccat gaggagaaa cgagatgct agtaaagaa gaattagga attggtgaa gaagaaaca gaagacgac agatttgac ttggaaaaa ccactgact gacattgtg attgtggga aacgctgac gatataatt atcgacgat tacgcagcc gactgcttt cccgattca tgttacaag aagtttccg tttcttgca ggcgacgag gattcgccg ttttggcag ggttggggg aatttaagg tcgaagact tatagacta atagagaac aaatatttt gaaacggcc gtaatcacg atgattttg ttgagtagc atggcgttg gcgttagaa gacgtacat ttgcaaagt cggccgatt ttgcaagac atactgtat tacatggat cgaatattc accgtgata ttctttttc gagatgttg ataaagtgg ttagcgtta gggttccaa aaatacttt acgaacgct tggtgctgg ctagatttt gtgattgta atggtgtct ctcattaat tttattgcc tctctttgt ggcgctggc gggattcaa gcatttaaa actatgcga acgctgcgg gctttgaga ccacttcgc gccatgtcc cgcatgcaa gggatgagg gttgtcgtt aatgcattg gtgcaagcg ataccgtct atcttcaat gtactgcta gtgtgctta atattttgg ttaattttt gctataatg ggtgtgcag ttattcgct ggtaaatac tttaagtgc gtggatgca aacaaaacc acactaagc tatgagatt atccctgat gttaacgcg tgtaaagct gagaattac acgtgggaa aactcccca atgaatttt gatcatgtg ggcaaagca tatttgtgc ttatttcaa gtggcgaca ttcaagggg tggattcaa attatgaac gatgctata gattctaga gacatcaac aaacaacca ataagagaa acaaacatt tatatgtat ttatacttc gtatttttt ataatattt ggatcgttt tttacactt aacttattc attggtgta atcatcgat aattttaac gaacaaaag aaaaagggt ggcgattca ttggcactg atgatgaca gacgcccag aagaaatat tacaatgcg atgaaaaaa atgggctct aagaaacca atgaaggcg ataccgagg ccgaggtgg agacctcaa gctatagtt ttcgaaata gtaacaaat aagaaattt gatatgata atcatgttg tttattgga ctgaacatg cttacgatg acaatggat cactaccaa cagaaagaa actttcacg aaagtcctc gattccctc aacatgatt ttcattgtt atctttagt accgagtgc ctcatgaaa atttttgcg ttacgttac cactacttt acggaacct tggaacctg ttcgattta gtggtagta atattatca attttgggt ttggttttg agtgacatc atagagaaa tatttcgtg tcaccgacg ttgttaagg gtggttcgt gtggccaaa gtagggcga gttctgcgt ttggtgaaa ggtgcaaag ggcattcgg acactgctg ttcgcacta gccatgtca ctgccggca ctgttcaac atctgtttg ttgctgttc ctggtcatg ttcattttt gccatcttc gggatgtcg ttcttcatg cacgtgaag gacaagagc ggacttgac gatgtctac aactttaaa acgtttggc cagtcgatg attctcctc ttccagatg tccacttcg gcgggctgg gatggagtc ctggatggt attatcaat gaagaagat tgtaaaccg ccaaacagt gagatcggc gagaccgga aattgcggg aattcgaca attggtatc gcattcctg ctcagctat ctcgtgatt tccttcctc atcgtcatc aacatgtac atcgctgtg attttggag aactactcc caggcgacg gaggatgtg caagagggt ctgaccgat gatgactac gacatgtac tacgagatc tggcagcag ttcgaccct gacggcacc cagtacatc cgctacgac caactgtcg gacttgttc gatgtgttg gagcctccg ctacagatc cacaagccc aacaagttc aaaatctac tccatggac attccgatc tgcaagggt gacatgatg ttttgtgtg gacattttg gacgcgcta acaaaagac ttcttcgct cgcaagggc aacgaaatc gaggaaact gctgaactt agtgaagtg caaccgaga cagaacgaa ccgggttac gaacaagtc agttcgacg ctttacaga cagcgcgaa gaatattgt gcgagactg atccagaat gcctggcgc aagtataga cagcgcact ggcggtgct ccagagcaa gtgaatgtc gatggggct gccggcgag ggcgatggg gaggtggag gtccaccaa acggccgtt ctggtcgag agcgacggc tttgtgacc aagaacggg cacaaggtt gtgctgcac agtcgcaca cccagcatc agctccaag tctgcggac gtctga
- >TC004749 RA
caagttctc tttttattt tcaggttag ttatgtttt tggccccca aaattgccc aatctgaat acgaggtat cgtgggggc ctaggcgcg aggagacat gtccgttgc ctctgacta ttcgtccga ggaggagct cagcctatt ccgtccatt cactcgcga gtctcttgc ggtcatcga gacgagaat tgcggaaga atatgctaa gcagaagga gctcgaaaa gaaaagagc tgaaggaga gattcggta tgaggacga ggacgaaga tgaaggtcc ccagcctga tccaactct ggagcaggg gctgccctt gccagtacg tcttcacag tacctttcc tccggaact ggccagtac accacttga agacatcga cccttttta cagtaacca aaggacttt tgtagtggt tagtaaagg caaggacat tttccggtt tagcgccac aaacgccct gtgggtact cgatccctt caatccgat ccgtcgagt ggccattta cattctagt gcatccttt gttttctct ttttatcat taccacaat cctcgttaa ctgtatcct tatgatcat gcccacaac acccaccgt agagtcaac cgaagtgat attcacagg catctacac gtttgaatc agctgtgaa ggtgatggc gagaggttt tatcctaca accgttcac ctaccttag agatgcatg gaactggct cgacttcgt agtcatagc tttagctta tgtcaccat gggtataga ccttgggaa tcttgctgc cttgagaac atttagggt actacgagc tctaaaaac tgttgccat tgtgccagg tttgaagac tatcgtggg agctgtaat agaatctgt aaaaaatct ccgagatgt gataatttt aacaatgtt ttccctgtc agtgttcgc attgatagg tttacaaat ttatatggg tgttttaac acaaaaatg tatcaaaga atttcctct ggatggctc ttgggggaa cctcaccca cgaaaactg ggagagatt tgttagcaa cgaatccaa ctggtattg ggatgacga aaaaggcga aaggcctct gtgtggaaa ctcctcagg atcgggaat gtgcaaaga gggttacgt ttgtttgca aggttacgg tgataaccc aaactatgg ttacaccag ctttgatac ttttggttg ggcattact ttccgcctt cagattaat gactcagga ttattggga gaatttata ccaactggt tttaagatc agccggtcc ttggcatat gctattttt tatcgtgat tatattttt gggttcgtt ttatttagt taacttgat cttggctat cgtcgctat gtcgtacga cgagttgca gaagaaagc ccaagatga ggaagaagc tgaagctaa cgctataag ggaagccga gcaaaaggc aaaggacaa gcaagatcg ggcggatgc gagggcagc ggcggccgt ggaggcggc cgaggcagc ggccgctgc cgcggcggc ggcggcagc agggtgtga caacgacca gtacatcat aaaatcgtc ttcggacta ctcgtgtca cagctatga gtcgttcgt ggggcaacc caacggtca agacgacaa caacaagga gaagattag catacgctc ggaaggatt agattccgt gagtgaaca gagaagagt tcctactaa tctgagcaa gatgcggaa agtcagcgc tgccagtct cagcctccc cggctctcc cttcaacct tcggcgagg gtcccgagg tagccacca gttcacctt gcggaacaa taggcgcat ggtggctcc gccgggcga tcgaaagcc cttggtgct gtcgaccta ccttgacgc ccaagagca cctccccta cgccgacga ctctaacgc cgtcacccc catgagcga ggagaacgg cgtcatggt cctgcccat gtactacac caacctggg atcacgtca ttcatcgta cacttcaca tgcctcacg gatgtcata cacttctca tggagattt gttaggcgg tttgggaag caatggcaa aatgatgac gaaggagag ccagctcag gaaccgatc gctgagaaa cggaccagc ggccgctcc cctaaacca caagtcaac gcgaggcgt ctacgatgg tcccacggg gcacatttg cgaatcaaa agttaagca aattgacaa tccttttat cgatagcaa tcagcgaca gacagtcgt ggacatgaa agatgttat ggtgctgaa cgacatcat cgagcaagc tgccgtgag cggaggcag tgaccgcgg agccaataa tgacgacga agaggaaga gccgacgtt cagggaacg cttcctggc ttatgccat caaaatcat tgacatgtt ttgcgtatg ggattgttg taaatgttg ggtcgttat tcagaagtt tgtcgccct tcttgtctt cgacccatt cgtcgagct ttttatcac gctgtgcat tgttgttaa caccttgtt tatggcatt ggatcacca cgacatgga caaggattt ggaaaaggc tttgaagag cggaaacta cttttttac tgcgacttt tatgattga agctacttt aaaattgat tgcaatgag tccgaaata ctactttca agaaggctg gaacatttt tgattttat tatagtggc gttatcgtt gctggagct aggactgga aggcgttca aggcttatc ggtcttgcg ttcatttcg actgttacg agtgttcaa actggccaa gtcgtggcc aactttaaa tttactcat ttccataat gggtagaac gatgggtgc tttaggaaa cttgacttt tgtgctctg tattataat atttatttt tgccgttat gggcatgca actgtttgg gaaaaatta cacagataa cgtggatcg gtttcccga ccacgagct cccccgctg gaacttcac cgacttcat gcattcgtt tatgatagt ttttcgcgt tttgtgtgg cgaatggat cgagtccat gtgggactg tatgctggt gggcgacgt ctcttgcat tccgttctt tctcgcaac tgtcgtgat tggcaatct tgtggtcct taatctttt cttagccct gcttttgag taactttgg atcttcgag tctgtcggc gccgactgc cgacaatga cacgaataa aatagcaga ggcttttga cagaatcgg ccgctttat cagatggat caaaggcaa tgttttaca tatcgcaaa actggtccg attcaagct caccaacca aatttccga ccagccgtc agatgtgag atgtgatgg tggtttaga tatctccgg agacgaaat tttagccga cggtataat taaaaaaag tcctaaaga ccgactgga agtaacaat tggcaccgg aatggatct taccgtgca tggtgatcc caaaagagg caaaaacaa tattaacaa aagcaaaac aatcggaaa ttcaattct ggaacatgg aatgtacat cggtcacct ggacgaaga cgaaattag taacaaatc gtacggtag tcacaaaaa tcgatttaa agaggaaag tcataaagg gagtgttga tatgttaga cgaccatga ggagaaacg agatgctag taaagaaga attaggaat tggtgaaga agaaacaga agacgacag atttgactt ggaaaaacc actgactga cattgtgat tgtgggaaa cgctgacga tataattat cgacgatta cgcagccga ctgctttcc cgattcatg ttacaagaa gtttccgtt tcttgcagg cgacgagga ttcgccgtt ttggcaggg ttgggggaa tttaaggtc gaagactta tagactaat agagaacaa atattttga aacggccgt aatcacgat gattttgtt gagtagcat ggcgttggc gttagaaga cgtacattt gcaaagtcg gccgatttt gcaagacat actgtatta catggatcg aatattcac cgtgatatt ctttttcga gatgttgat aaagtggtt agcgttagg gttccaaaa atactttac gaacgcttg gtgctggct agattttgt gattgtaat ggtgtctct cattaattt tattgcctc tctttgtgg cgctggcgg gattcaagc atttaaaac tatgcgaac gctgcgggc tttgagacc acttcgcgc catgtcccg catgcaagg gatgagggt tgtcgttaa tgcattggt gcaagcgat accgtctat cttcaatgt actgctagt gtgcttaat attttggtt aatttttgc tataatggg tgtgcagtt attcgctgg taaatactt taagtgcgt ggatgcaaa caaaaccac actaagcta tgagattat ccctgatgt taacgcgtg taaagctga gaattacac gtgggaaaa ctccccaat gaattttga tcatgtggg caaagcata tttgtgctt atttcaagt ggcgacatt caaggggtg gattcaaat tatgaacga tgctataga ttctagaga catcaacaa acaaccaat aagagaaac aaacattta tatgtattt atacttcgt attttttat aatatttgg atcgttttt tacacttaa cttattcat tggtgtaat catcgataa ttttaacga acaaaagaa aaagggtgg cgattcatt ggcactgat gatgacaga cgcccagaa gaaatatta caatgcgat gaaaaaaat gggctctaa gaaaccaat gaaggcgat accgaggcc gaggtggag acctcaagc tatagtttt cgaaatagt aacaaataa gaaatttga tatgataat catgttgtt tattggact gaacatgct tacgatgac aatggatca ctaccaaca gaaagaaac tttcacgaa agtcctcga ttccctcaa catgatttt cattgttat ctttagtac cgagtgcct catgaaaat ttttgcgtt acgttacca ctactttac ggaaccttg gaacctgtt cgatttagt ggtagtaat attatcaat tttgggttt ggttttgag tgacatcat agagaaata tttcgtgtc accgacgtt gttaagggt ggttcgtgt ggccaaagt agggcgagt tctgcgttt ggtgaaagg tgcaaaggg cattcggac actgctgtt cgcactagc catgtcact gccggcact gttcaacat ctgtttgtt gctgttcct ggtcatgtt catttttgc catcttcgg gatgtcgtt cttcatgca cgtgaagga caagagcgg acttgacga tgtctacaa ctttaaaac gtttggcca gtcgatgat tctcctctt ccagatgtc cacttcggc gggctggga tggagtcct ggatggtat tatcaatga agaagattg taaaccgcc aaacagtga gatcggcga gaccggaaa ttgcgggaa ttcgacaat tggtatcgc attcctgct cagctatct cgtgatttc cttcctcat cgtcatcaa catgtacat cgctgtgat tttggagaa ctactccca ggcgacgga ggatgtgca agagggtct gaccgatga tgactacga catgtacta cgagatctg gcagcagtt cgaccctga cggcaccca gtacatccg ctacgacca actgtcgga cttgttcga tgtgttgga gcctccgct acagatcca caagcccaa caagttcaa aatctactc catggacat tccgatctg caagggtga catgatgtt ttgtgtgga cattttgga cgcgctaac aaaagactt cttcgctcg caagggcaa cgaaatcga ggaaactgc tgaacttag tgaagtgca accgagaca gaacgaacc gggttacga acaagtcag ttcgacgct ttacagaca gcgcgaaga atattgtgc gagactgat ccagaatgc ctggcgcaa gtatagaca gcgcactgg cggtgctcc agagcaagt gaatgtcga tggggctgc cggcgaggg cgatgggga ggtggaggt ccaccaaac ggccgttct ggtcgagag cgacggctt tgtgaccaa gaacgggca caaggttgt gctgcacag tcgcacacc cagcatcag ctccaagtc tgcggacgt ctgagagcg gccccgccc ctcccctac ctcgccgta gcctgagcg gctcaggag gcacgtctt tttcgtctc ggctgtacg ccagactac cctcactgt tacatcgaa ttccataag aattgcgtt cattgcgtt aagaccgag actaaaatc acgtcgact caagtctgt gctgtgtag tttggtgga gaaggtatc gtataacta ggtgtcaac tgcttttgc tgctcatat cgctatcgt actatttaa ctaatggtt acttaactg aaatccaca ccctcaacc agacatagt gtatgaaca gtgaattta accactaac tagtcctct attttcaca cagatcgtt cgattagaa tgactgtcg taattctag tcgaatgca atgtggcaa aatgtccca gcatttggt gccacatgt gaaaatacc gacactagt ccgagacag ttgatgtag tcactgaat cctgttaat tcgaaatgt agagttttg gtttctgtt acccctctg tgacacact ttgacttaa tgggacgac agttattat aattttgca gtaaatcct gtaaatagt cttgttgaa accggaaca ggtagaaca aagcttggc gcgagccga gcatatcat cctgagtct attagcctt aaaatgccg ccaaagcaa tcatttaga ctaaaaatt tagcaaatt gttatttgg gagcaaccg ttcccaagt ggctcccaa aactcccca cgattttaa ggataaaga ttcagggac ttccaagtc aaattcgcg aaaagtgta tgattttga tccgattcg ggcacaaat ttattgaat cgaggaata cacgtttcg aggattggc cgcaaatgt tatattcta aatgggccc gaatcatca gttatttgt gaaattatg gtgctttct tcaattgct aatcttcaa gctatcttc acatgctac atggaccgc attcaaggg aggggcaaa tgggttaaa atttgcaaa aatacacaa atttttgta cgcctcatg taaattcgt atgtgtgac atttttgtg gagcaacag gcacacggc agttttgta attagttag ctgaaatag tggttgata tttcattta ataaacacc aaagcattt taagacgtg atggcacgg ttgtacaaa ttatacttc acaaaattg tacattatg tagggctgt ttatacgtc atcctaaat tttgtatca tggctggac cattaacta agtaataac gtgtccatc attttaaaa tgtgtggaa aaatgtcac gcataccct tgcaacaac tttattgat aatgtaact agggcaggg gtgcagcct cttgctcga agttacgcc acgctgtta cattttgtt aattattaa taatattaa ttgattgaa taattatta ttgttgtta aagttttat atgaatatt acggaattg gttagtgtt tgtttgacg tggtttatt gcgtatgtt attatcatt gtcatcatt atattcccg taatctttc ttggtccca caagccatt ataatatat gatactcaa cctg
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