The invention provides a straw rotting edible fungus liquid strain medium, a preparation method and an application. The straw rotting edible fungus liquid strain medium comprises, in weight percent, 0.5%-1.0% of glucose, 0.5%-1.5% of straw rotting edible fungus sporophore enzymatic hydrolysate, 5.0%-10.0% of edible fungus secondary fermentation culture material extract liquid, 1.5%-2.5% of oil sunflower cake boiling liquid, 0.05%-0.15% of yeast extract and the balance water. The liquid strain medium is rich in nutrient component and reasonable in composition, the growth requirement of straw rotting edible fungi is sufficiently met, so that mycelia can rapidly germinate and grow to occupy growth space, sundry fungus resistance is enhanced, activity is strong, stability is good, the myceliaare slow in aging, and the preparation success rate of a straw rotting edible fungus liquid strain is remarkably increased.