Molecular sex determination with DNA markers is a gender determination on genetic level. With their help gender can be determined in very young plants or even seeds. Sex determination of plants before its phenotipic expression is useful in plant breeding processes. Induction of male flowers on dioecious female plants is a process in which with the help of growth regulators we influence the expression of sex. The practical purpose of the method of male-flower induction on dioecious female plants is to enable self-polination or crossing plants that are genetically female. In this way we can obtain pure lines and we also obtain feminized seeds. The goals of the master 's thesis were to test the markers for cannabis sex determination SCAR119, SCAR323 and MADC2, to optimize the PCR mixture and the temperatures of primers annealing for sex determination in cannabis and to test the effect of growth regulators (GA3, STS), and their various treatments on the expression of sex on dioecious female plants of medical cannabis. The most efficient treatment was the use of undiluted STS after which we obtained the highest number of male inflorescences and flowers. The marker SCAR119 proved to be the most effective marker for gender determination. The best technique for obtaining results in accordance with phenotypic observations was the technique with the use of isolated DNA and its amplification with the SCAR119 marker. We also optimized a sex determination protocol for i.e. direct PCR, where preliminary DNA isolation is not required, and we tested it on 200 hemp plants. The success for gender determination was 75%.