Shivani Gupta-Week 6- Reddy Lab

Shivani Gupta-Week 6-Reddy Lab

                Unfortunately, our colonies for the BirA-YY1 fusion did not grow. We determined that the reason must be that we didn’t add enough DNA in our ligation reaction. We did the ligation over, and grew the colonies on amp plates. Luckily, about 25 colonies grew on the BirA-YY1 plates. I then did the mini-preps over using these colonies and then digested the BirA-YY1 with Xmal. After doing the restriction digest verification, we found out that only one of our 10 samples had the correct band length and we could send to sequencing.

                We then picked 10 more colonies from our plates and repeated the cloning procedure by first doing mini-preps and then the restriction digest verification. Unfortunately, I didn’t have enough DNA in my samples and had too much RNA, so I couldn’t see the bands when I imaged my gel for the digest verification. We think that it was because we may have lost some of our DNA during the mini-preps. Next week we are going to grow new BirA-YY1 colonies on our plates, and hopefully we will be successful.

                Thankfully, one of the sequences for the BirA-L2B matched the original construct, so I have successfully made a fusion protein!

                On Thursday, Dr. Crider came to visit my lab. Jen and Dr. Reddy came to lunch with Dr. Crider and I, and we ended up eating lunch in the newer part of the hospital. The cafe was so crowded, we ended up sitting at a kids table, but it was still a great experience!