Solid phase microextraction (SPME) is a modern technique to isolate volatile compounds and it can be an alternative to traditional methods. It was successfully applied to analyse different plant fruit and food products. The authors of this paper focused their attention on the aroma of transgenic cucumber modified using a gene of thaumatin II. Until now, no results have been published of any studies dealing with the evaluation of the effect of this type of genetic modification on the volatile profile of raw material. The objective of this study was to select parameters of the SPME technique for evaluating the aroma of cucumber. The parameter optimization of the SPME technique comprised the selection of: type of fibre, temperature, extraction time, and salting out the sample. Owing to the application of the SPME technique, it was possible to identify volatile compounds responsible for the aroma of cucumber. Moreover, it was found that the contents of volatile compounds in the fruit of transgenic lines and in the non-modified control line differed.