Aim: The present study aimed to carry out DNA extraction and polymerase chain reaction (PCR) amplification to isolate the physiological and phenotypic traits of Candida albicans and Candida dubliniensis in denture wearer patients with and without denture-induced stomatitis. Materials and methods: A total sample size of 160 participants were divided into two equal groups (80 each), patients in the study group having 40 males and 40 females with Newton type II denture stomatitis, and in the control group, healthy 40 males and 40 females those who wear complete denture were selected. All the samples were collected from the hard palate with a sterile swab and inoculated on CHROM agar plate; samples that displayed dark green colored colonies were selected for DNA extraction. DNA isolation was done on agarose gel using electrophoresis. Biorad gene identification was used. Strands depicting the presence of DNA in particular samples were identified, and further standardization of the procedure was done. PCR amplification was done using Candida species-specific primer, preset to the hyphal wall of the protein 1 gene with the CRR forward and reverse primers, under strict standard conditions with reverse transcriptase technique. Results: Results showed that prevalence of C. albicans was more in females with denture stomatitis which was 67.50% than in males, i.e., 52.50%, and prevalence of C. dubliniensis was found in one female and in one male who were having denture stomatitis and it was not isolated from patients without denture stomatitis. Statistical analyses were performed using the Chi-square test. Conclusion: Denture stomatitis is the most common problem faced by long-term denture wearers, with C. albicans as one of the causative organisms. However, recent findings show an emerging pathogenic yeast species, C. dubliniensis, which was isolated from denture-induced stomatitis candidates in the present study, which is closely related to the C. albicans species. The identification of candidal strains causing denture stomatitis with DNA extraction and PCR amplification and its management by determination of its susceptibility to antifungals may improve the treatment outcome of the same. Clinical significance: Candidiasis is the most frequently seen mucocutaneous infection of the oral cavity especially in denture wearers. It is caused mainly by the genus Candida. C. dubliniensis is phenotypically similar but genotypically different from C. albicans. This affects the treatment outcome drastically as there is enough literature suggesting resistance to the common antifungal drugs. Hence, drugs like fungus-specific calcineurin inhibitors should also be considered in resistant patients. Therefore, DNA identification of Candida genus plays a major role in deciding the treatment outcome.