Gene editing in yeast cells using the CRISPR/Cas9 system
Zhang Tengyu and Melissa A. Mefford
Morehead State University, Morehead KY USA
Background
CRISPR/Cas9 is a recently developed
technology to edit the genome of any
organism. The CRISPR/Cas9 system was
initially discovered in bacteria, where it
functions as an essential defense system
against viral invasion by bacteriophage.
CRISPR (clustered regularly interspersed
palindromic region) DNA is transcribed
and processed into guide RNAs that target
complementary DNA sequences while the
Cas9 protein cleaves the DNA.
Figure 1. Principle of CRISPR/Cas9 gene
editing. Figure from https://www.vox.com/2018/
7/23/17594864/crispr-cas9-gene-editing
RESEARCH POSTER
PRESENTATION
DESIGN © 2015
Project Overview
I will use CRISPR/Cas9 to edit three
genes in the yeast Saccharomyces
cerevisiae: ADE2, TLC1 and PXR1.
Successful editing of ADE2, a gene
involved in adenine synthesis, will cause
yeast to accumulate red pigment and turn
pink. Successful deletion of TLC1, the
RNA component of the telomerase
enzyme, will cause yeast to senescence.
Successful editing PXR1, a gene involved
in rRNA and snoRNA processing, should
result in slow growth.
Experimental Plan
Current Progress
Figure 2. Building pCAS with TLC1 and PXR1
gRNAs individually. Candidate bacterial colonies
containing modified plasmids.
Future Directions
1. Assess the efficiency CRISPR gene
editing in yeast.
2. Further study the role of PXR1 in
telomere maintenance.
3. Design the experiment with
CRISPR technology to analyze the
other function of ADE2.
Acknowledgements
Thank you to KBRIN New Faculty Start Up Award to Dr.
Melissa A. Mefford for funding(NIGMS PZOGM103436) and
Elieen Sylves from the State University of New York Buffalo
for sharing reagents.