Saccharomyces cerevisiae has been known for its applications in a wide range of industries such as food and beverages, pharmaceuticals, bioethanol and feed industries. Mitr Phol Innovation and Research Center has successfully isolated and developed a few Saccharomyces cerevisiae strains for their ethanol plants located in four different regions of Thailand. Industrial ethanol production in Thailand is typically conducted without a strictly aseptic bioreactor using molasses as the feedstock. In order to increase ethanol production and its efficiency, the yeast strains require the ability to tolerate ethanol concentrations higher than 10\% (ethanol v/v) and stresses from molasses. Since the conventional method to isolate yeast strains from natural resources is laborious, random and costly, it is necessary to use an alternative method using bioinformatics and systems biology for better understanding of the genetic-phenotypic relationship that helps to discover target genes for yeast improvement. It was found that our isolated strains (MP11, MP15) show high ethanol production compared with the commercial strain Angel Super Alcohol (Angel Yeast, Co. Ltd.). Therefore, we selected 10 important genes (ADH1, ADH3, ADH4, ADH5, HXT1, HXT2, HXT3, HXT4, TPI1 and SUC2) based on previous publications to clone and sequence candidate genes of Mitr Phol's yeast strains. After that, we applied comparative sequence analysis among yeast strains to identify DNA variation in genes that are relevant to ethanol production. From the results, the most variation of DNA sequence was found in a primary enzyme for ethanol production (ADH1) of about 12\%. In addition, the variation of ADH1 was also found at substrate binding sites among our isolated and commercial strains. This result implies that genetic variation among yeast strains has an effect on ethanol production especially on the key enzyme in the ethanol pathway. In order to implement yeast strains with the specific trait to improve ethanol production, genetic information of each yeast strain is required to further apply this strategy for stress tolerance traits.