roller phenotype from RNAi?

Model Organism: Worms Scientific Discussion
1

Hi All,

Does anyone know whether there are any genes that produce a good roller phenotype with feeding RNAi? This is for a class demonstration, so I am not picky about which gene it is. From Wormbase, it seems that RNAi on many rol genes produces either no phenotype or lethality. RNAi for rol-4 says you see rollers but also Dpys and some lethality. I would like to get the most robust and clean roller phenotype possible so the students don’t get confused. I have the Ahringer RNAi library, so I should have what I need in the freezer.

Thanks,

Tim

2

Tim,

I believe that the roller phenotypes arise from dominant missense mutations in some of the cuticle collagens. These mutations lead to a strangely kinked collagen subunit the is twisted and leads to a twisted cuticle. As far as I know, decreasing any of the cuticle collagens with RNAi will not lead to a roller phenotype. This is true of rol-4 (sqt-3) and rol-6.

On the other hand, a strong DPY phenotype can be quite distinctive. I am not sure which cuticle protein would give you the strongest phenotype.

Janet

3

rol-5 RNAi works well, while rol-6 RNAi does not (su1006 is a gain of function allele and the sequence is used as an co-injection marker).

4

I agree that rol-6 RNAi is ineffective at producting a Rol phenotype, but I should point out that the canonical allele of rol-6, e187 (https://cgcdb.msi.umn.edu/strain.php?id=7072), is a recessive loss-of-function mutation. It is true that we all use the stronger, neomorphic, gain-of-function allele for co-injection markers. But logically, one would expect a Rol phenotype from rol-6 RNAi, but I am guessing the efficiency of knocking down this gene is poor.

Steve

5

Using an RNAi hypersensitive strain might be good to get loss of function rollers out. As Steve points out, that is the rol-6 lf phenotype.

Another possibility is the Sqt genes, where heterozygotes are Rol but homozygotes are Sqt? Presumably sqt RNAi could induce a Rol phenotype.

My recommendation for a class would be unc-22: RNAi causes the coolest Unc phenotype there is.

6

e187 is recessive for Rol, but is missense, so it could easily be recessive gain-of-function. A better case can be made by reference to n1268 and n1270, which are identical W36ochre stop mutations and so are likely null mutations (n1178 is a late amber, and so less obviously a null).