Gaykar Amruta B.
PTGS in plants:Discovery of Jorgensen in 1990   AIM: upregulate activity of chsA      Expected:more pigments       Observ...
RNA interference – gene silencing by double-strandedRNA:The 2006 Nobel Prize for Physiology or Medicine    CURRENT SCIENCE...
Mechanism of RNAi   Two major pathways    involved     miRNA       Control of endogenous       gene expression    siRNA...
Geminivirus induced gene silencing in plants                               (Ramachandran et. al., 2005)
Transgene directed gene silencing in plans     Sense     Antisense     Sense- Antisense                            (Wat...
Introduction   Four major tomato-infecting tospoviruses        TSWV        Groundnut ringspot virus (GRSV)        Tomat...
Constructs used in gene silencing
Results
Suppression of RNA silencing by viral proteins
(Miguel, 2007)
SGS3-mediated generation of siRNAs
Genome organization of RCNMV
RNA1 is essential for suppressing RNAi
Introduction     BBTV, a circular single-stranded DNA virus     Genus- Babuvirus     Family- Nanoviridae     Multi-seg...
Materials and Methods Plasmids and Agrobacterium      The ORF fragments B1, B3, B4, B5 and B6 of BBTV       DNA1, DNA3, ...
Transcription cassettes for Agroinfiltration
   BBTV B3 , B4 and B5    enhance the pathogenicity    of chimeric PVX                                N. benthamiana leav...
Western blot analysis      Accumulation of PVX CP in N. benthamiana leaves
N. benthamiana leaves infectedwith chimeric PVX under UVlight at 5 dpi                   GFP accumulation detected by west...
Results:14 dpi       BBTV B4, but not B3 protein, suppresses        systemic gene silencing in newly emerged        leaves
Viruses fight back against rn ai technology
Viruses fight back against rn ai technology
Viruses fight back against rn ai technology
Viruses fight back against rn ai technology
Viruses fight back against rn ai technology
Viruses fight back against rn ai technology
Viruses fight back against rn ai technology
Viruses fight back against rn ai technology
Viruses fight back against rn ai technology
Viruses fight back against rn ai technology
Viruses fight back against rn ai technology
Viruses fight back against rn ai technology
Viruses fight back against rn ai technology
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Viruses fight back against rn ai technology

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RNA silencing refers to related processes of post-transcriptional control of gene expression found in plants, animals and fungi. A unifying feature of RNA silencing is that it mediates sequence-specific degradation of target transcripts, recruiting RNA molecules of 21–23 nucleotides as specificity determinants. In higher plants, RNA silencing serves as an adaptive, antiviral defence system, which is transmitted systemically in response to localized virus challenge. Plant viruses have elaborated a variety of counter-defensive measures to overcome the host silencing response. One of these strategies is to produce proteins that target the cell autonomous or signalling steps of RNA silencing. It is not known whether a similar antiviral mechanism also operates in animal cells

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Viruses fight back against rn ai technology

  1. 1. Gaykar Amruta B.
  2. 2. PTGS in plants:Discovery of Jorgensen in 1990 AIM: upregulate activity of chsA Expected:more pigments Observed:white sectors Loss of mRNAs of both endo-and transgene (Etienne et al., 2006)
  3. 3. RNA interference – gene silencing by double-strandedRNA:The 2006 Nobel Prize for Physiology or Medicine CURRENT SCIENCE, VOL. 91, NO. 11, 10 DECEMBER 2006
  4. 4. Mechanism of RNAi Two major pathways involved miRNA Control of endogenous gene expression siRNA Defense against viruses and transposons (Lindbo et al., 2005)
  5. 5. Geminivirus induced gene silencing in plants (Ramachandran et. al., 2005)
  6. 6. Transgene directed gene silencing in plans  Sense  Antisense  Sense- Antisense (Waterhouse and Helliwell ,2003)
  7. 7. Introduction Four major tomato-infecting tospoviruses  TSWV  Groundnut ringspot virus (GRSV)  Tomato chlorotic spot virus (TCSV) and  Watermelon silver mottle virus (WSMoV), in a single small chimaeric hairpin (hp) RNA construct
  8. 8. Constructs used in gene silencing
  9. 9. Results
  10. 10. Suppression of RNA silencing by viral proteins
  11. 11. (Miguel, 2007)
  12. 12. SGS3-mediated generation of siRNAs
  13. 13. Genome organization of RCNMV
  14. 14. RNA1 is essential for suppressing RNAi
  15. 15. Introduction  BBTV, a circular single-stranded DNA virus  Genus- Babuvirus  Family- Nanoviridae  Multi-segmented genome  B3 and B4,encoded by DNA3 and DNA4 of virus(BBTV), exhibit RNA silencing suppression activity
  16. 16. Materials and Methods Plasmids and Agrobacterium  The ORF fragments B1, B3, B4, B5 and B6 of BBTV DNA1, DNA3, DNA4, DNA5, DNA6 were obtained by PCR using primers  The obtained fragments were inserted into pND108 Agroinfiltration and GFP imaging Western blot analysis
  17. 17. Transcription cassettes for Agroinfiltration
  18. 18.  BBTV B3 , B4 and B5 enhance the pathogenicity of chimeric PVX N. benthamiana leaves infected with chimeric PVX at 14 days post infiltration (dpi)
  19. 19. Western blot analysis Accumulation of PVX CP in N. benthamiana leaves
  20. 20. N. benthamiana leaves infectedwith chimeric PVX under UVlight at 5 dpi GFP accumulation detected by western blot analysis with anantibody specific for GFP at 5 dpi B3 and B4 reverse established RNA silencing of GFP
  21. 21. Results:14 dpi  BBTV B4, but not B3 protein, suppresses systemic gene silencing in newly emerged leaves

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