Chapter 18, Problem 34P

One potential strategy for gene therapy to correct the effects of dominant gain-of-function mutations is to express small interfering RNAs (siRNAs) that will cause degradation of mRNA produced by the dominant mutant allele. The siRNAs can be delivered to patient cells by a synthetic gene encoding a hairpin RNA that will be processed by Dicer and used to target the RISC complex to the mutant allele mRNA (review Figs. 17.32 and 17.33).

a.	A major problem with this gene therapy strategy is designing an siRNA that will prevent the expression of the mutant allele specifically and not the normal allele. Explain why this is a problem.
b.	Specificity for the mutant allele is a particular problem in designing an siRNA therapy gene for Huntington disease. Explain this issue and suggest a possible solution that would allow you to use RNAi.
c.	Another potential strategy to correct the mutant Huntington disease allele is to cut the DNA in the middle of the repeat tract. Exonucleases present in the nucleus would then degrade many of the repeats prior to repair by NHEJ. Could you use CRISPR/Cas9 technology to correct Huntington disease mutations with this approach? (Hint: See Problems 28 and 29.)