Interactions of soil moisture with plant root systems are very important for plant growth. For non-invasive determination of volumetric soil moisture in a rhizobox, a microwave system based on transmittance of electromagnetic waves in the microwave frequency range was developed using microstrip patch antennas. Vector Network Analyzers (VNAs) were used to measure the S-parameters at frequency ranges close to 5 GHz. A transmission system with microstrip patch antennas was developed. The result of this attenuation is in the frequency domain. The antennae were designed as resonant microstrip antennae. The antennae were placed on both sides of a rhizobox, which allowed non-invasive measuring soil moisture in the box. The attenuation (S21(dB)) was used to measure the effect of temperature, and different types of soil; as well as sensitivity, reproducibility and repeatability of the system. In this work we present quantitative results of soil moisture in rhizobox. The microwave technique, using microstrip patch antennas, is a reliable and accurate system, and showed very promising potential applications for rhizobox-based investigations of root performance.