English
Construction of CRISPR/Cas9 mediated gene editing system of Hypsizygus marmoreus
Beomgyu Park, Hyeon-Su Ro*
Department of Bio & Medical Big Data and Research Institute of Life Sciences, Gyeongsang National University, Republic of Korea
*Email: rohyeon@gnu.ac.kr
Growing concerns about environmental impact, dietary restrictions such as veganism, and health issues have increased interest in alternative foods. Mushrooms have attracted attention due to their β-glucans that support immunity, mycoproteins, and distinctive flavor and texture. In Asia, various mushroom species have long been consumed, and in recent years, Western markets have expanded consumption beyond Agaricus bisporus, driving global demand for greater varietal diversity. Hypsizygus marmoreus, an edible species widely consumed in Asia—particularly in Japan—is now increasingly cultivated in Korea and China. It possesses high nutritional value, containing abundant proteins, vitamins, and minerals, and has strong market potential. To enable molecular genetic studies, we established a CRISPR/Cas9 gene-editing system for H. marmoreus. Preassembled Cas9–gRNA ribonucleoprotein (RNP) complexes and hygromycin resistance gene expression fragments were introduced into protoplasts via PEG-mediated transformation. Screening through 5-FOA, Hygromycin-containing medium will confirm the disruption of pyrG gene and the expression of hygromycin resistance gene. These results establish a reference system for gene editing in H. marmoreus, providing a foundation for gene function analysis and variety development.
