Of the 2 parent strains and 6 single-spore isolates (SSIs) used in this study, the SSIs BBSR-007 and BBSR-002 of the culinary-medicinal straw mushroom Volvariella volvacea exhibited superior growth characteristics on different growth media. These also took less time until first harvest (days after spawning), gave higher numbers of fruiting bodies per unit weight of substrate, and provided a higher mushroom yield on composted substrate. The fruiting body weight of isolate BBSR-007 was significantly higher compared to BBSR-002. On pasteurized paddy straw, the SSI BBSR-007 had a higher mushroom yield than BBSR-002. The contents of dry matter, protein, and other elements (sodium, potassium, and calcium) were on par in both the parent strains and different SSIs, except SSI OE-55-08, which had highest dry matter, protein content, and potassium/-to-odium ratio. In amplified ribosomal DNA restriction analysis, the SSIs did not show any distinctness, whereas in amplified fragment length polymorphism, the SSI OE-55-08 of parent strain OE-55 formed a separate clade; of 4 SSIs of strain OE-274, the SSI BBSR-003 formed a separate clade than other SSIs. The genetically distinct SSI OE-55-08 produced the highest numbers of fruit bodies per unit weight of substrate and exhibited the highest amounts of dry matter and protein in its fruit bodies. Another slightly distinct SSI, BBSR-003, formed white aerial mycelia on growth substrate, and its fruit bodies exhibited the highest levels of sodium and calcium.