Post about The Lab

In this post I am going to talk about lab stuff, and try to explain some of it. So here it goes. During the first three weeks or so, I spent my time isolating RNA from xenograft tumor and normal samples. The xenografts were tricky but do-able, however the normal samples were a huge pain in my... I managed to succeed finally after many tries. Only then, to take the samples out of the freezer a couple of days later to find that they were all degraded. So I was very thankful when another fellow in the lab, Andy, (thank you Andy) was able to successfully isolate normal RNA and give some to me. From there I was able to proceed in doing many RT-PCRs. That is a Reverse Transcription - Polymerase Chain Reaction, which is converting the isolated RNA into cDNA and then amplifying certain sequences of it to eventually compare expression levels between normal and tumor samples. So that's what I have been doing for the past 3 weeks now. Here is a picture from my lab meeting presentation of the hippo pathway that I will explain below.
Click for hippo size
The Hippo Pathway is a growth regulation pathway, first discovered in flies, but also present in mammals, which when activated stops the growth of certain organs. Animals who lack this pathway have large overgrown body parts, and look like hippos, which is where the name came from. The picture shows the known components. The five that I circled are the genes that I have been looking at. Next picture.
Click for make bigger here
This picture shows the results after running the PCR product on an agarose gel. Each gene has its own row, and each sample its own column. GAPDH was used as a control, but it didn't work out so well because all the bands were supposed to be the same strength. Pay attention to the difference between the average normal sample, OD Mix, and the rest. Especially for Yap-1, which the tumor samples are over expressing by far. More pictures.
Engorgio (harry potter joke)
This picture has the other two genes, and GAPDH as well. Same story as before although the first ones were better. Because the here the no template control started acting up again and I have no idea where that band came from, it's supposed to be empty. And sample 552 just disappeared in some places, which hasn't been helping things. So that's where I am right now in the lab. My next thing to do is to use a new control other than GAPDH, in order to try to get more equal expression levels. I hope I did a good job of explaining. You can ask questions in the comments section below. I turned it on it should work. This is my last week in the lab so there might only be one more post after this one ;(