Initiation and growth of short cracks in a nickel-based single crystal were studied by carrying out in-situ fatigue experiments within a scanning electron microscope (SEM). Specimens with two different crystallographic orientations, i.e., [001] and [111], were tested under load-controlled tension fatigue in vacuum. Slip-caused crack initiation was identified at room temperature while initiation of a mode-I crack was observed at 650 °C. Slip traces continuously developed ahead of the crack tip once initiated and acted as nuclei for early-stage crack growth at both room and high temperature (650 °C). These slip traces were caused by accumulated shear deformation of activated octahedral slip systems, which were specifically identified by analysing the surface slip traces and crack-propagation planes. The crack-growth rates were evaluated against stress intensity factor range, revealing the anomaly of slip-controlled short-crack growth. The effects of crystallographic orientations and temperature on fatigue crack growth were subsequently analysed and discussed, including the influence of microstructural features such as carbides and pores.