The enzyme myrosinase (EC 126.96.36.199.147) is present in specialised myrosin cells and forms part of the glucosinolate–myrosinase system, also known as ‘the mustard oil bomb’, which has an important role in the defence system of cruciferous plants against insect pests. Transgenic Brassica napus MINELESS have been produced by transgenic ablation of myrosin cells. This prompted us to investigate the importance of myrosin cells in plant–aphid interactions. In order to study this, we challenged transgenic MINELESS and wild-type cultivar Westar seedlings with the aphids Brevicoryne brassicae (a specialist) and Myzus persicae (a generalist). Our study included aphid free-choice and aphid fecundity experiments. Data from these experiments showed that B. brassicae prefers wild-type seedlings and M. persicae prefers MINELESS. B. brassicae and M. persicae showed significant variation in establishment on plants regardless of whether they were wild type or MINELESS and also differed significantly in affecting plant parts. Myrosinase activity in MINELESS control seedlings was 83.6% lower than the wild-type control seedlings. Infestation with either of the two aphid species induced myrosinase levels in both wild-type and MINELESS seedlings. Infestation with M. persicae reduced the concentration of most glucosinolates while B. brassicae had the opposite effect. B. brassicae enhanced the formation of glucosinolate hydrolysis products both in wild-type and MINELESS seedlings. However, M. persicae decreased All ITC but increased 3,4ETBut NIT in wild-type seedlings. Taken together, the investigation shows that the presence of myrosin cells affects the preference of generalist and specialist aphid species for Brassica napus plants.