Isolation and Regeneration of Wheat Mesophyll Protoplasts: Using a Short Protocol

Plant protoplasts serve as valuable tools for studying the fundamental properties and functions of cells. For example, protoplasts have been used to analyze cell organelles, the cell cycle, and cell wall deposition. Another application involves protoplast fusion to create somaclonal hybrids and facilitate genetic modifications. Additionally, plant protoplasts are successfully utilized in experiments aimed at mitigating heavy metal toxicity, plant stress, and perception responses, as well as robust expression screening systems for recalcitrant species. Protoplasts from various plant species have become essential in genome editing technologies, with their application established in numerous laboratories today. Here, we present a reproducible and concise method for isolating wheat mesophyll protoplasts from two spring varieties, Canon and Alderon, derived from in vitro seedlings. A cell counter and fluorescent staining were employed to determine the concentration and viability of freshly isolated wheat protoplasts. A modified culture medium initiated the first cell divisions of one wheat genotype within 8 days after the protoplasts were placed in the culture.