White Leghorn (WL) breed is homozygous for an allele on the α2C-AR gene while the Red Junglefowl (RJF) is mostly heterozygous for that. The gene is also hypermethylated in WL. The gene codes for the α2C -adrenergic receptor, which plays various roles including regulation of neurotransmitter release from adrenergic neurons in the central nervous system and sympathetic nerves The aim of this study was to investigate the effects of the variation of α2C-AR gene on the chickens’ behaviour. Various behavioural tests mostly associated with fear and stress was conducted in progeny of an F9 generation of the advanced intercross line (AIL) between RJF and WL, selected for homozygosity of the alternative alleles on the α2C-AR gene. In the emergence test, the stress effect on both poking and total exit latency was significantly higher in WL genotype females in comparison to RJF genotype females (effect on head poking time: WL -70.62 ± 18.603 versus RJF 15.63 ± 29.069; effect on total exit time WL -72.14 ± 20.582 RJF 5.64 ± 30.140). In the aerial predator test RJF genotype birds showed significantly more agitated behaviours after the predator exposure in comparison to WL genotype birds (RJF 27.65 ± 0.700 versus WL 25.09 ± 0.915) Although we found differences in behaviour of individuals with WL genotype, more research is needed to find out how the variation on the ADRA2C gene has played a role in domestication of layer chicken.