The two weeks I spent at the Fred Hutch Explorers Program were such a transformative and welcoming experience. The schedule during the past 10 days was packed with labs, many tours across different campuses, guest speakers, lectures, as well as interactive group activities. For me, the highlight of my experience was the CRISPR lab.
Picture of a poster introducing the topic of CRISPR and steps.
During this lab we had to conduct PCR to make many copies of the B2M gene, which contains a mutation known to increase a person’s risk of cancer. PCR reagents are temperature sensitive, so we had to keep them in sterile environments and keep the reagents on ice during waiting times. In the CRISPR lab we cut the B2M PCR product using CRISPR/Cas9 and two guide RNAs. The specific guide RNAs that I chose for this lab had a fragment size of 26-45 and 472-492.
The procedure for this lab involved labeling the two guide RNAs (gRNA) and then binding them to the Cas9 and incubating them together. Next, we prepared the tubes of the gRNAs by adding reagents of PCR-grade water, 10x Cas9 buffer, the gRNA and Cas9 nuclease. After mixing the tubes and incubating them, we set up the Cas9 reaction by adding RNA and waited before degrading any remaining Cas9 enzyme. The challenging part of this lab was the degradation of the Cas9 enzyme, as it required adding a very precise amount of Proteinase K to both tubes. This could have led to error as the micropipette was not precise in extracting exactly 1 ul of Proteinase K. However, after centrifuging both tubes and flicking them, we were able to extract the exact amount.
Furthermore, during the CRISPR lab we were required to make gels. We did this by mixing 15 ml of 1% agarose into each casting tray of the clear gels, and then waited for the agarose to solidify before placing the gel plate into the buffer chamber. When the gel tray was placed back into the gel box, we measured 140 ml of the 0.5X TAE and poured it into the tanks. The final steps were to prepare the Cas9.gRNA samples for gel electrophoresis and the B2M PCR-product for gel electrophoresis before loading the DNA ladder and samples into the wells. Once the DNA migrated down about 75% of the gel, we were able to record our results.
The gel electrophoresis revealed that our PCR amplification of the B2M gene was successful, showing clear bands of the expected size. It also revealed that the CRISPR/Cas9 system effectively targeted and attached to the B2M gene at the specific locations of the gRNAs. This lab impacted me because it required the skill set to precisely measure and work with hands on techniques, as well as exploring new technology in the lab.
Picture of accurate results of the CRISPR lab (grading down on the left).
Overall, my experience at the Explorers Program was not only interactive and informative, yet it also confirmed to me that I love working in labs and researching. Additionally, everyone that I met was kind and passionate about science, making this experience wholesome and fun.
Picture of most of the girls from second session of the Explorers Program. Picture of me (left) and my friend, Vivian (right).
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