I wanted to discuss something - I'm working on a report where we've isolated kallikrein-4 RNA from prostate cancer cells, made cDNA, amplified by PCR, and analyzed on a 1,5% agarose gel. I pasted the results above.
Sorry about the Norwegian in the text, basically we expected a 700bp fragment for KLK4 (which we see in lane 2 from a KLK4 expressing plasmid).
In lane 5, you can see the KLK4 cDNA we amplified by PCR. Now for what I'm wondering about ... we see a lot of noise compared to the plasmid KLK4.
I'm wondering why this could be, and was hoping you guys had any idea. I'm an undergraduate with a good deal of experience with gels, but I haven't done exactly this before.
- We see another clear band other than the 700 bp fragment + some other noise
- is the other band a result of alternate splicing? I suppose the mRNA is already spliced since cDNA doesn't contain introns. Do you think the noise could be because of some cancer-specific process?
- I'm worried about the noise we see over the whole lane in well number 5 - is this simply because of PCR-byproducts?
Thanks. I just wanted to mention that I don't really need this for my report, it's somewhat extracurricular, but I'm just personally curious since this was a lot of fun. :)