Sunday, 19 January 2014

Lab fail

Since Christmas I've been cursing the fact that I've been sent a fly with a deleted gene only to discover a diagnostic PCR for said deletion reveals the gene is still there. I thought the fly may be on some "subtle" (the dreaded ultrabithorax*) balancer and that I was checking a heterozygous fly. I checked with the lab that sent the fly and they believed it was homozygous. Cue a lot of grumbling about people sending dodgy stocks.

The problem was that the deletion mutant would be really useful so I thought I'd design another set of primers because it just may be the case that I had got an off-target band that was roughly the same size as the band I was expecting. Desperate, I know.

The thing is when I got around to designing the primer I made sure I had the full info on the deletion as well. Eyeballing it, I thought "the deletion looks a lot bigger than the whole gene". That's when I realised I was comparing the genomic DNA of the deletion with the cDNA (the DNA that only encodes the protein). That's when it dawned on me that the primers I had designed for the diagnostic test were cDNA specific. The 1.5kb band I was expecting would be around 33kb from the genomic DNA. Never going to work with the PCR protocol I have!

So thanks to my ineptitude the deletion may well still be there. I've ordered the appropriate primers and will know by the end of the week whether it's real or not. Fingers crossed it is - I'd rather be an idiot than have to try and make a mutant from scratch.

*Ultrabithorax may sound like a really cool Decepticon but in reality is a really hard to distinguish phenotypical marker in Drosophila that only those gifted with "the sight" can easily recognise. The phenotype is present on the haltere of the fly which is like a tiny secondary wing that is used for manoeuvrability.

A) Wild Type  B-D) ubx130/Wild Type.  Figure taken from

You need to keep in mind that the haltere is very small to begin with.

Picture taken from

Basically I hate it. I'd love it a lot more if the marker was as penetrant as the homozygous mutant I'd love it a whole lot more because the phenotype is super cool.

Picture taken from

Da Vinci Dragonfly!  

EDIT: Fans of "Jonathan Strange and Mr Norrell" should appreciate the footnote to text body ratio in this post :P

No comments:

Post a Comment