The Hybridization Barrier between Herbaceous Medicago sativa and Woody M. arborea Is Weakened by Reproductive Abnormalities in M. sativa Seed Parents

Historically, crosses between Medicago sativa (alfalfa) and M. arborea with alfalfa as the seed parent failed, as did crosses using M. arborea as the seed parent. Thus, a reproductive barrier kept the two species isolated until early in this century. The breakthrough came when alfalfa seed parents were identified in Wisconsin USA and Queensland AU that produced partial hybrids (hereafter hybrids). The hybrids were obtained by making large numbers of crosses on selected alfalfa parents. This was the first level of weakening the crossing barrier as reported in Plants in 2013. Further weakening of the barrier is reported herein whereby more hybrids were obtained with fewer crosses. This was accomplished by pedigree selection for new alfalfa seed parents and by using a product of the first hybrids called Alborea. New alfalfa seed parents were crossed with M. arborea, and Alborea parents were backcrossed to M. arborea. Hybrid plants were produced with fewer crosses in both cases. These hybrids, like the first hybrids, have mostly alfalfa traits but also have traits from M. arborea. It was theorized early on that the alfalfa component could be explained by 2n eggs in the alfalfa parents that were fertilized by normal n gametes from M. arborea. Evidence that the Wisconsin alfalfa and Alborea seed parents did in fact produce 2n eggs was reported in Plants in 2022. Moreover, they produced 2n eggs at approximately the same frequency that they produced hybrids. As reported herein, Alborea parents produced the highest frequency of hybrids and thus had the weakest barrier. Importantly, they also have the highest frequency of 2n eggs. It was determined that alfalfa and Alborea parents that produce 2n eggs and hybrids, also produce 2n pollen. In effect, an experiment was undertaken in reverse showing that 2n pollen could be used to screen for plants that produce hybrids. In the thousands of crosses made over the years, fertilization of normal n eggs in alfalfa parents always failed. Normal meiosis appears to be the main barrier to producing interspecific hybrids in our case. Fertilization of abnormal 2n eggs ensures sufficient alfalfa genetic material to continue embryogenesis. Evidently, the meiotic abnormality of 2n eggs is the major factor that weakens the crossing barrier.