Salinity, an important abiotic stress, affects arable land worldwide, reducing crop growth, yield and quality. Lettuce ( Lactuca sativa L.), which is very sensitive to salt stress, is one of the crops affected by soil salinity and salt (NaCl) stress also affects lettuce cultivation. Different plant growth regulators (salicylic acid (SA), polyamines, glycine betaine (GB) and melatonin (MEL)) are used to reduce the harmful effects of soil salinity. In this study, the effects of NaCl and MEL treatments on physiological parameters (cell membrane damage (%), leaf relative water content (RWC%), chlorophyll amount), DNA methylation and some NaCl tolerance genes ( SOS1, SOS2, AKT1, NHX1, HKT1) have examined in two different lettuce genotypes (Yedikule and Ivanka). When the effect of NaCl stress on physiological parameters was examined, it was detected that this stress increased cell membrane damage and decreased chlorophyll content and RWC %. The effects of MEL applications together with NaCl stress on these parameters were positive. In the gene expression analysis results, it was determined that SOS1, SOS2, AKT1, NHX1 and HKT1 gene expressions increased in both lettuce genotypes and decreased in MEL applications. On the flip side, according to methylation sensitive amplification polymorphism (MSAP) findings, while a decrease in methylation level was determined in NaCl stress compared to the control in both genotypes. Methylation levels increased compared to NaCl stress in MEL applications with NaCl stress, but this rate was still lower than the control. According to the results obtained, it was concluded that exogenous MEL application in lettuce genotypes is important in response to NaCl stress.