1. Technologies: RNAi and Variants, Nuclease Variants
Label or no label…. I say it is genetically modified
Wayne Parrott
Department of Crop and Soil Sciences
Institute for Plant Breeding, Genetics & Genomics

2. RNA interference Unifying concept that explains Gene silencing
Resistance to viruses
Many natural traits
Della Vedova et al 2005; Genetics 170:
Burpee
Tuteja et al., 2004; Plant Cell 16: 2

3. Low glutelin rice Green maize stems 3
C2/C C-Idf/C2
Green maize stems 3
Kusaba et al Plant Cell 15:

4. Use of RNAi in agriculture4

5. Silencing pathways + siRNA High levels of RNA Overexpression
Co-suppression
Sense suppression
MicroRNA
Hairpins
Antisense RNA +
siRNA

6. Natural silencing caused by genomic rearrangements when not due to microRNA genes
C-Idf C2
Della Vedova et al Genetics 170:

7. DNA sequence to be edited
Genome editing Based on cutting DNA & allowing natural repair of the cut
DNA sequence to be edited
DNA sequences
Cut DNA at site to be edited
Provide a copy of DNA needed to replace gap
Desired DNA added at designated spot
Option 1:
1. Insert genes into specific sites
2. Trait correction/conversion
Replace one allele with another
ATTGGGATCAAGC
ATCGGGATCTAGC

8. DNA sequence to be edited
Genome editing Based on cutting DNA & allowing natural repair of the cut
DNA sequence to be edited
DNA sequences
Cut DNA at site to be edited
Option 1:
Option 2:
DNA gap filled with random DNA
(Gene permanently turned off)
Why Agrobacterium, gene gun, & transposons and recombination can leave a footprint
Provide a copy of DNA needed to replace gap
Desired DNA added at designated spot

9. Making the cut at a specific site Zinc Finger Nucleases
Zinc Fingers:
Proteins designed to recognize 3 base-pairs
Nuclease
Moore et. al,  PLoS ONE. 2012;7:e37877

10. Making the cut at a specific site Zinc Finger Nucleases & TALENs
Moore et. al,  PLoS ONE. 2012;7:e37877

11. Handle locks DNA guide into place
DNA needed find target site
Handle locks DNA guide into place
DNA cutting enzyme
DNA sequence to be edited
SCIENCE 341: AUGUST 2013

12. Hybrid systems

13. Non pathogen-derived resistance
Glyma07g14530 A
STOP
STP
STOP G
Can recreate resistance
RNAi
CRISPR/Cas9

14. Cis-genes From plants from the same family
Rpi-vnt1.1 gene Solanum venturii engineered into S. tuberosum
A type of type of gene common in all plants
Would never be regulated if crossed in conventionally
Marker genes
Not all traits/genes need them
Segregate out
Cut out
Consider them inert material

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