Unfavorable environmental conditions during planting can reduce seed germination and hinder seedling growth. To address this issue, manufacturers are exploring innovative and cost-effective methods, such as cold plasma discharge. This simple, low-cost, and efficient physical technique induces significant biological responses in seeds and plants without the use of traditional, environmentally hazardous chemicals. This study investigated the impact of dielectric barrier discharge (DBD) plasma and plasma-activated water (PAW), produced by gliding arc plasma, on the germination and seedling growth of My344 cotton seeds. The seeds were pre-treated with 80 W of DBD plasma for 0, 1, 2, and 3 minutes, and subsequently soaked for 90 minutes in PAW with varying pH levels of 5.82, 3.88, 3.63, and 3.38. The results showed that plasma treatment positively influenced seed germination and seedling growth. The highest germination percentage (98.89%) was observed with 1 minute of DBD treatment, followed by PAW priming at pH levels of 3.63 and 3.38. Additionally, a 3-minute DBD treatment followed by soaking in PAW with a pH of 3.63 led to significant increases in stem length (76.76%), root length (48.77%), and wet weight (76.44%). Furthermore, it was observed that the electrical conductivity of the seeds in all groups decreased significantly with increased PAW acidity. The physical and chemical effects of cold DBD plasma on the seed surface, as well as changes in hydrophilicity, were further examined using scanning electron microscopy (SEM), Fourier-transform infrared spectroscopy (FTIR), and water contact angle imaging.