A new towing taxi-out departure mode has become the preferred choice to replace the traditional departure mode of civil aircraft due to its obvious advantages of low energy consumption, low emissions, and high efficiency. However, the inertia may lead to potential lateral instability and even “Jack-knifing”, as the mass of the system is concentrated in the rear. In this paper, the lateral instability mechanism and influencing factors of the civil aircraft towing taxi-out system were studied using computer virtual simulation technology and taking Boeing 737–400 and AM210 tractors as research objects. The results show that the “Jack-knifing” phenomenon was caused by the lateral instability of the system when the sliding speed of the system was more than 15 km/h, and the system was accompanied by the lateral slip condition. Furthermore, the safety zone, side-show zone and jack-knifing zone of the system were defined in terms of towing speed and steering angle. Additionally, a towing safety zone was created under different airport track conditions by analyzing the lateral stability of the system. The conclusion of this study may provide a theoretical basis and reference for the safe application of the new taxi-out departure mode.