No day is the same working in the Poss Lab. Every day consists of new experiments to carry out, new things to learn, and new questions to ask.
The first two weeks of lab largely consisted of shadowing my secondary mentor and learning basic protocols. After gaining experience and comfort in conducting PCR reactions, gel electrophoresis, heart extractions, embryo injections, cloning, and cryosectioning (just to name a few), I have really enjoyed gaining independence in my work.
My project looks at 4 different genes and they’re currently all at different stages in my experiments. I have already ran in situ hybridization for one gene while I’m still at step 1 in trying to isolate the segment of DNA I want for another gene. Thus, every day I continue working on my project, whether it’s the final step for transcribing my RNA probe or trying to troubleshoot a PCR for the tenth time. My days will usually involve running multiple PCR reactions and gels. If my gel runs properly, I can extract the DNA band and purify it. Then, I clone the DNA into a plasmid and transform it in bacteria. After the bacteria grows, I set up a colony PCR to verify the DNA segment and send the plasmid for DNA sequencing. If the DNA sequence looks good, I inoculate more bacteria, extract the DNA with a midi-prep, linearize the plasmid, purify, and transcribe (if all goes smoothly) with many PCR’s and gels in between in order to double check each step. Usually, a step won’t work correctly and I have to try to figure out what the problem is. Troubleshooting a step may take days, so I repeat a step many times, changing one thing at a time in order to determine what is wrong.
Depending on the day, I may also practice embryo injections which involve setting up zebrafish matings the day before. I collect the embryos, prepare the injection mixture, and inject hundreds of zebrafish embryos. I may also need to make cDNA for my PCR which involves dissecting zebrafish hearts, extracting the RNA, and reverse transcribing it to make cDNA. I am usually conducting many experiments at once. While I’m waiting for my PCR, I may be embedding hearts in tissue freezing medium or screening zebrafish for mutants. Some days I am busy going back and forth from the lab to the fish room. While other days I have more waiting time when I can read some papers.
My routine largely consists of the same protocols, but working every day in lab is still exciting. Although creating an RNA probe for each gene involves the same steps, each gene behaves differently. Thus, applying each step to each gene creates different results, so I never know what I’m going to get. Sometimes, things don’t work out so it can be challenging but I enjoy trying to solve the puzzle. I have definitely learned that science takes time but I am excited to discover what it will reveal.