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Trehalose is a common energy reservoir, and its accumulation results in osmotic protection. This sugar can accumulate through its synthesis or slow degradation of the reservoir by trehalase enzymes. <i>Saccharomyces cerevisiae</i> contains two neutral trehalases, <i>NTH1</i> and <i>NTH2</i>, responsible for 75% and 25% of the enzymatic metabolism. We were interested in the loss-of-function of both enzymes with CRISPR/Cas9. The later <i>NTH2</i> was of great importance since it is responsible for minor metabolic degradation of this sugar. It was believed that losing its functionality results in limited osmotic protection. We constructed an osmotolerant superior yeast capable of growing in 0.85 M NaCl after independent <i>nth2</i><i>1271_1272delTA</i> mutation by CRISPR/Cas9 technology, compared with <i>nth1</i> <i>893_894insT</i> and wild type. We suggest that this yeast model could give clues to breeding commercial yeast resulting in non-GMO salinity-tolerant strains.