Due to the ability to synthesize a highly potent neurotoxin, Clostridium botulinum poses an important risk to food safety. Foodborne botulism is a neuroparalytic disease caused by the ingestion of neurotoxins produced by Clostridium botulinum. Botulism commonly occurs as a result of consuming canned foods. In this study, the aim was to investigate the presence of C. botulinum and detect the genes producing A, B, E, and F toxins in C. botulinum isolates in homemade and commercial canned foods using the rapid, sensitive, and reliable Real-Time PCR technique. A total of 81 canned samples were collected, including 51 commercial and 30 homemade cans. The canned samples were initially enriched in tryptone peptone glucose yeast (TPGY) medium. Subsequently, DNA isolation was performed using the foodproof® StarPrep Two Kit. After the isolation process, Real-Time PCR was conducted using the foodproof® Clostridium botulinum Detection LyoKit and 5’ Nuclease (Biotecon, R60240) kit. Only 2 samples tested positive among all the samples. Both positive samples were identified to contain neurotoxin type A. They were both commercial canned foods—one was a mixed vegetable garnish, and the other was roasted red pepper. Both positive samples were packaged in glass containers. Consequently, it was concluded that commercially available canned foods in Turkey could still pose a potential risk to public health due to the presence of C. botulinum. Therefore, it was recommended to perform quality control analyses through food inspections in companies involved in canned food production and sales.