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| − | <li><a href="https://2015.igem.org/">iGEM</a></li>
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| − | <h1>Practices</h1>
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| − | <h2>RNAi: Off-Target and Non-Target Effects</h2>
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| − | <p>The use of RNA interference in pesticide does not come without the possibility of negative effects or errors in what RNAi sequences target. Ideally, the RNAi we manufacture should only target the specific sequence that causes virulence in Fusarium graminearum. Carrying out the proper function in this manner is an on-target effect. Due to the sequence similarities across species and within the genome of the target pest, there could be unwanted effects. Two types of these effects are off-target and non-target.
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| − | <p>Off-target effects are when the processed RNAi silences a part of the genome that is downstream or upstream of the target sequence, or only part of the target sequence. RNAi technologies have been found to have lack of sequence specificity (Jackson and Linsley, 2010). Therefore, a virulence factor may not be out, allowing F. graminearum to continue infecting crops. A 100% sequence match is often required for the RNAi to work (Subba Reddy Palli, 2014), so any slight sequence change or mutation could render the RNAi sequence as unable to disrupt the target sequence. It is therefore important to know what the exact sequences are that will cause death in the organism when down regulated. Just because a 100% match is required for the RNAi to work doesn’t mean that it can’t interact with and effect another area within the genome that is slightly different, impairing its function as well. Further testing of the RNAi spray would be required to determine the exact effects regarding the probability of off-target effects, but it has been shown that using a lower concentration of interfering RNAs reduces the amount of off-target effects within the genome (Dharmacon, 2014).</p>
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| − | <p>Non-target effects are when the RNAi down regulates a gene that is not in the intended target species. Due to the short length of interfering RNAs, there is more opportunity for sequence similarities between species. Therefore two different species could be equally likely targets of the RNAi. This means that the spray could influence the growth and function of other pests, pollinators and crops and even cause death in these organisms. Similar to off-target effects, it is possible that due to lack of specificity a non-target species could have a sequence with a completely different function than the target interact with the RNAi. It is therefore important to determine what species in these agricultural communities could be effected by the chosen sequence, and what sequence similarities exist between species that are closely related to the target. Off-target and non-target effects could potentially occur at the same time. For example, a non-target species could have short lengths of sequence similarity with F. graminearum and so it’s genome was disrupted by the RNAi, but due to RNAi’s lack of specificity there was off-target silencing downstream of the matching sequence within that organism’s genome.
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| − | <h2>References</h2>
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| − | <li>Dharmacon. 2014. Off-Target Effects: Disturbing the Silence of RNA interference (RNAi). 1-4.
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| − | <li>Jackson A. and P. Linsley. 2010. Recognizing and avoiding siRNA off-target effects for target identification and therapeutic use. Nature. Vol. 9: 57-67.
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| − | <li>Subba Reddy Palli. 2014. RNA interference in Colorado potato beetle: steps toward development of dsRNA as a commercial insecticide. COIS. 3:1-8.
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