As a trial for the development of a new starter culture for yogurt products, more than two hundred lactic acid bacteria strains were isolated from raw milk and healthy human feces. The strains that showed excellent growth and acid production ability in the 10% skim milk media were selected and identified as Lactobacillus casei through the API carbohydrate fermentation pattern and 16S rDNA sequence analysis. L. casei CU2604 was further investigated for its physiological characteristics as a starter culture compared with a commercial strain. The CU2604 strain showed good acid production and growth characteristics in milk, which were comparable to those of the L. casei Shirota strain. Despite the fact that both these strains displayed the same sugar fermenting pattern and PFGE band pattern, and had similar growth characteristic in milk, L. casei CU2604 exhibited different fatty acid composition in the cell wall, showed more tolerance to bile and to pH, and presented better growth inhibition activity against pathogenic bacteria. Based on these results, the L. casei CU2604 strain holds great promise for use as a novel and efficient starter culture in the production of yogurt. Additional studies on the probiotic characteristics of this strain are currently being conducted.