Biology Reference
In-Depth Information
Table 14.6: Successes and Concerns with RNAi in Insects.
Successes
RNAi allows scientists to evaluate the function of genes in many insects. Species that have had their genomes
sequenced readily allow effective ds RNAs to be designed for many different genes.
RNAi can be used in cell cultures or in living organisms. Systemic RNAi in living organisms is most relevant
for determining the function of genes involved in development, reproduction, behavior, immunology, and
other aspects of insect biology. Systemic RNAi involves the spread of RNAi effects from cell to cell and
tissue to tissue in the living insect.
By 2010, RNAi had been used in species of arthropods in the orders Orthoptera, Blattaria, Isoptera,
Hemiptera, Coleoptera, Neuroptera, Hymenoptera, Lepidoptera, and Diptera, as well as in some mites
and ticks (Acari), indicating RNAi can provide functional genomics data for many arthropods.
Concerns
RNAi may result in off-target effects, leading to knockdown of genes with structural similarity to the delivered
ds RNA.
RNAi may inhibit gene expression, but
rarely
eliminates it. It is not a knockout, but a
knockdown
procedure
that can be transient. If the effects are too transient, a phenotype may not be detected.
Arthropods vary in their ability to develop a systemic response to RNAi, probably for several reasons
including: efficient degradation of alien ds RNA by some species; deficient amplification and spread of
the ds RNA signal; the tissue is not permeable to the ds RNAs; the gene counteracts RNAi by increasing
transcription rates; the target mRNA is too transient or is protected from RNases. It appears that “less-
derived species” are more likely to express systemic responses.
The mechanisms underlying amplification and spreading of the RNAi signal are not fully understood in
arthropods.
Methods for delivering ds RNA to arthropods vary, including injection, feeding, electroporation, or soaking.
The amount of ds RNA required to elicit a response also varies.
For RNAi to function in insect control in crop plants, the insect must take up the ds RNA of a target gene
through feeding; the ds RNA should pass through the gut lumen into the gut cells; and, if the target gene is
expressed outside the gut, the silencing signal will have to spread (systemic RNAi). The nucleotide sequence of
the ds RNA should not function in other insects (nontarget effects). If the silencing effect is transient, effective
control may not be achieved. The optimal life stage of the insect to be targeted should be determined.
If crops are produced that are resistant to major pest insects, resistance to RNAi could develop. Questions
include: should RNAi function continuously and in all plant parts at high concentrations? Should RNAi
genes be “stacked” to reduce the likelihood of resistance in the pests?
Other issues relate to the risk assessment of RNAi plants; are there risks unique to RNAi mechanisms?
Derived from
Belles (2010), Huvenne and Smagghe (2010), and Center for Environmental Risk Assessment (2011).
may be transient, which could induce resistance in the target pest. One solution to
delay resistance in the pest would involve stacking target genes (pyramiding).
14.6 Methods to Deliver Exogenous Nucleic Acids into Arthropod
Tissues
A variety of methods have been evaluated for delivering genes and vectors into
arthropods to achieve transformation (
Table 14.3
). Current methods include
