Biochemistry Unveiled: A Comprehensive PowerPoint Presentation

Hi guys, welcome to my video. So, today I'm going to show you what it's like to work in a biochem lab. So, I've been working on protein expression, purification and also crystallization. I'm crystallizing my proteins at the moment. I'm really excited to see the structure of it later on. I've been filming a lot in the lab and I'm very excited to show you guys how it's like to work here, experiments that I've done, which hopefully you guys would find interesting and also you could get to know more about what daily life is in the lab. First thing I do when I arrive is to make a brief lab plan which would include today's objectives and experimental steps. Today, I need to grow some cell cultures so I'll be making some agar plates. We've picked some colonies and now we're growing more of those cells which have taken up the plasmids and we will grow more of them to produce our protein. Here for this experiment, I've used IPTG to induce expression. This foamy layer indicates a really good growth rate, so now I would have to see whether there's enough cells by measuring the OD value. What I'm doing here is a step called strip and recharge which is in preparation for protein purification. The nickel column has been cleaned up by the step strip and recharge earlier in the video and I'm using it for the second round of protein purification. Before the purification, I also ran some gels to confirm whether the protein was present. This is SDS-PAGE which denatures the protein and I also add a blue stain to stain the gel which makes visualization easier. So I've just finished purifying my protein and I've ran my proteins in the SDS-PAGE gel so I'm waiting for it to separate and hopefully I'll see like a nice pattern when I go back. I'm outside just for a short walk and I'm gonna go back. Yo, are you ready? I'm already filled in mate. So the gel has been stained and it shows that I have so much protein that it basically exploded the well. This is really good news because this means I get to play around with lots of my protein, I can test them with different conditions, I can test with a lot of crystallization conditions. So here this is protein dialysis which is another round of protein purification. Pool sizes of the bag are a lot smaller than the proteins which means that the protein will be staying inside the bag and other components that we don't want will be filtered out of the bag. The final round of protein purification will be size exclusion chromatography by the actor. The actor separates the sample into different fractional columns. I'll be running another gel to see what happens after purification by SEC. Now it's time for the final round of purification and I'm going to inject the protein sample into the actor in order to run the SEC column. So that's why that one's called H Red Sea. After purifying the protein I've collected them into fractions so I'm just testing out which fractions actually contain the purified protein sample. Here I'm making a buffer in preparation for tomorrow's experiment so I can start straight away tomorrow morning when I come back. So since I've finished making the protein now I will be determining its structure and I'm doing it by x-ray crystallography. I will be making a separate video on how I do crystallography, how I make crystals and if you're interested in how to make protein crystals please go watch my video. It's a very fun process and I'd love to share it with you guys. That's it for today. Thank you so much for watching and I hope you liked it. Please let me know in the comments what contents you guys would like to see and I'm really excited to see you guys again.