Spinal instrumentation with pedicle screw placement (PSP) is an important surgical technique for spinal diseases. Accurate screw trajectory is a prerequisite for PSP. Ultrasound (US) imaging with robot-assisted system forms a non-radiative alternative to provide precise screw trajectory. This study reports on the development and assessment of US navigation for this application. A robot-assisted US reconstruction was proposed and an automatic CT-to-US registration algorithm was investigated, allowing the registration of screw trajectories. Experiments were conducted on ex-vivo lamb spines to evaluate system performance.In total, 72 screw trajectories are measured, displaying an average position accuracy of 2.80 ± 1.14 mm and orientation accuracy of 1.38 ± 0.61°.The experimental results demonstrate the feasibility of proposed US system. This work, although restricted to laboratory settings, encourages further exploration of the potential of this technology in clinical practice.