The present disclosure provides a display substrate, a manufacturing method thereof and a display device, and relates to the field of display technology. The display substrate includes a base substrate, and a driving circuitry layer and a light-emitting unit arranged on the base substrate. The driving circuitry layer includes a first gate metal layer, and a pattern of the first gate metal layer includes a gate line. The first gate metal layer includes a first conductive layer with resistivity less than a first threshold and a Young's modulus less than a second threshold, and a first conductive protection layer arranged at a side of the first conductive layer away from the base substrate.