Using TD‐PCR, we generated trnL‐L‐F sequences from 84 samples among the 16 putative Lomariopsis populations in Taiwan. Excluding the 15 samples found to belong to other genera, we identified 69 Lomariopsis gametophytes. In total, we collected 122 (ribbon‐shaped) fern gametophytes from the tree fern trunk, and 84% (102/122) of them were identified as V. junghuhnii using its taxon‐ specific primer set. In addition, we also performed TD‐PCR on all of the gametophytes using the universal primer trnL‐L‐F set, confirming that all pre‐treated samples could be amplified well using this technique. This indicates that the samples were fresh enough and contained limited secondary metabolite concentrations, which can inhibit PCR reactions. Using present‐day DNA tools, the identification of fern gametophytes in the field is more accessible. To improve these tools, TD‐PCR strategies can be incorporated not only to facilitate DNA‐based identification, but also to explore new research directions for the investigation of fern gametophytes in the field. As the cases here have demonstrated, TD‐PCR requires no DNA extraction and makes gametophyte identification possible using a single PCR step. Notably, this technique works well for the gametophytes of both ferns and bryophytes. Time, funding, labor, and research materials for gametophyte DNA identification can thus be directed to other advanced purposes. We are also looking forward to the further development of this molecular ecology technique and to improving the efficiency and reliability of the DNA‐based identification of fern gametophytes. Most importantly, with advances in fern biology and ecology, this featured methodology reminds us that the mystery of fern gametophytes in the field is still waiting to be uncovered.