There is a relative lack of basic information about early diverging species of the genus Medicago that, for the most part, were formerly considered to be in the genus Trigonella. Species boundaries are not always clear, for example, the most recent treatment of the genus Medicago submerged four previously recognized species into Medicago monantha, a widely distributed species in the Middle East. These species are recognized on the basis of morphological characters such as fruit number, shape, length and areole shape and size, but species identification is still challenging and further clarification of species boundaries is needed. There is also a lack of cytogenetic information. Some of the relatively few published chromosome numbers, e.g. 2n=28, and 44, differ from those of the rest of the genus, which are mostly 2n=16 or multiples thereof, although seven species are 2n=14. As part of a larger study of the genome and chromosome number evolution in the genus Medicago, we obtained genome size data using flow cytometry for 44 accessions of 14 currently recognized early diverging species, with a focus on Medicago monantha. Chromosome numbers were obtained using standard cytological methods. Our chromosome number data confirm a chromosome number of 2n=16 for M. brachycarpa (genome size of 1.33 pg), and M. monspeliaca (1.88 pg), and 2n=28 for M. polyceratia (2.77 pg) and give new numbers for some species; 2n=16 for M. biflora (2.7 pg), and a previously unknown chromosome number for these early diverging species of 2n=14 for Medicago fischeriana (~1.35 pg). Interestingly, our data support the hypothesis that there are at least two entities within M. monantha as currently recognized that differ in chromosome number and genome size; two accessions had chromosome numbers of 2n=26 and 30 with corresponding genome sizes of 2.68 and 2.85 pg and three other accessions had chromosome numbers 2n=36,44, and another 44 with genome sizes of 3.94, 3.89, and 4.04 pg. There are also some significant morphological differences between these two entities, such as fruit length and areole area. These data lead to both further clarification of the relationships of early diverging Medicago and help build a platform for more in-depth research concerning the evolution of chromosome number and genome size within Medicago.
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