Calcium ion (Ca) is a universal second messenger involved in regulating diverse processes in animals, plants, and fungi. The low-affinity calcium uptake system (LACS) participates in acquiring Ca from extracellular environments under high extracellular Ca concentration. Unlike most fungi, which encode only one protein (FIG1) for LACS, nematode-trapping fungi (NTF) encode two related proteins. AoFIG_2, the NTF-specific LACS component encoded by adhesive network-trap forming Arthrobotrys oligospora, was shown to be required for conidiation and trap formation. We characterized the role of DhFIG_2, an AoFIG_2 ortholog encoded by knob-trap forming Dactylellina haptotyla, in growth and development to expand our understanding of the role of LACS in NTF. Because repeated attempts to disrupt DhFIG_2 failed, knocking down the expression of DhFIG_2 via RNA interference (RNAi) was used to study its function. RNAi of DhFIG_2 significantly decreased its expression, severely reduced conidiation and trap formation, and affected vegetative growth and stress responses, suggesting that this component of LACS is crucial for trap formation and conidiation in NTF. Our study demonstrated the utility of RNAi assisted by ATMT for studying gene function in D. haptotyla.