Date

1-4-2017 12:00 AM

Major

Genetics

Department

Genetics, Development, and Cell Biology

College

College of Agriculture and Life Sciences

Project Advisor

Maura McGrail

Project Advisor's Department

Genetics, Development and Cell Biology

Description

Mutations in DNA mismatch-repair (MMR) genes have been observed in many types of human cancers, including malignant brain tumors. The main goal of this experiment was to study the contribution of DNA-repair towards malignancy of cells, with an objective to determine whether disruption of the MMR pathway will result in a change in the number of dividing cells in the brain. To study this interaction, I disrupted the MSH6 gene (MutS Homolog 6: a key DNA MMR gene commonly mutated in brain cancers), in a zebrafish model for pediatric brain cancer, which is deficient for the rb1 (Retinoblastoma) tumor suppressor. I employed the CRISPR-Cas9 gene engineering system to target frameshift mutations in the MSH6 gene, in zebrafish embryos. I embedded and sectioned the brains of the 5-day-old mutant larvae, and labeled the sections with markers for proliferation, etc., through immunohistochemistry. I then imaged the stained sections using confocal microscopy, and counted the number of proliferating cells in the brains. There was no statistically significant difference in proliferation between the injected and uninjected larvae. The next step would be to simultaneously target multiple MMR genes including MSH2, which may have compensated for the loss of MSH6 function in this case.

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Apr 1st, 12:00 AM

Studying The Role Of DNA Mismatch Repair In Brain Cancer Malignancy

Mutations in DNA mismatch-repair (MMR) genes have been observed in many types of human cancers, including malignant brain tumors. The main goal of this experiment was to study the contribution of DNA-repair towards malignancy of cells, with an objective to determine whether disruption of the MMR pathway will result in a change in the number of dividing cells in the brain. To study this interaction, I disrupted the MSH6 gene (MutS Homolog 6: a key DNA MMR gene commonly mutated in brain cancers), in a zebrafish model for pediatric brain cancer, which is deficient for the rb1 (Retinoblastoma) tumor suppressor. I employed the CRISPR-Cas9 gene engineering system to target frameshift mutations in the MSH6 gene, in zebrafish embryos. I embedded and sectioned the brains of the 5-day-old mutant larvae, and labeled the sections with markers for proliferation, etc., through immunohistochemistry. I then imaged the stained sections using confocal microscopy, and counted the number of proliferating cells in the brains. There was no statistically significant difference in proliferation between the injected and uninjected larvae. The next step would be to simultaneously target multiple MMR genes including MSH2, which may have compensated for the loss of MSH6 function in this case.