Arbuscular mycorrhizal fungi (AMF) are a group of soil microorganisms that establish symbiosis with most terrestrial plant species. “Root trap culture” has generally been used to isolate a single regenerated spore to establish a monospecific line of native AMF. Roots can be co-colonized with multiple AMF species; however, only a small portion of AMF within roots sporulate, and they do so only under certain conditions. Recent studies have proven that young (<2 mm) thalli of the liverwort Marchantia paleacea harbor monospecific AMF, and can be used as a vegetative inoculant line. When M. paleacea buds are co-cultured with field-obtained roots, young AMF-infected thalli via rhizoids and form arbuscules after 18 days post-sowing. Ribosomal DNA sequencing of AMF-colonized thalli (mycotalli) reveals that they harbor phylogenetically diverse AMF; however, new buds planted around transplanted mycotalli show evidence of colonization by phylogenetically uniform Rhizophagus species. It should be noted that mycotalli can also be used as inoculum. The results of this study suggest that young thalli of M. paleacea allow us to potentially isolate monospecific AMF from soil in a spore-independent manner.