Selection activities in the frame of the ongoing breeding programs led to a reduction of the diversity in genetic materials and are considered as a bottleneck in crop evolution after the domestication process. This assumption is accompanied by the idea that many alleles useful for breeding could have been lost in the selection process. Therefore, in the last few years a growing interest has focused on analysing genetic diversity in landrace collections in cereal crops.

In some cases, these studies were aimed at characterizing a limited number of important genotypes, such as those traditionally grown by farmers in particular areas. The genetic analyses, carried out on a proper number of individuals for each accession, often showed a certain degree of heterogeneity in each landrace. As an example, Mangini et al. [37] carried out a phenotypic and molecular analysis of three durum and one common wheat Italian landrace population, and the SNP characterization revealed different haplotypes within each landrace, indicating a genetic structure based on a mixture of genotypes. In other cases, landraces were maintained as inbred lines, and analysis on very large collections became possible. These analyses were often focused on panels of landraces with a specific geographical origin, as in the case of durum or common wheat landraces from Ethiopia [38,39], Sicily [40], Morocco [41], Iran [42], Palestine and Israel [43], Pakistan [44], Turkey [45] and Mexico [46]; barley from Nepal [47], the Canary Islands [48], Tunisia [49,50] and Jordan [51]; oat from Poland [52] and Spain [53]; rice from Pakistan [54] and India [55] and millets from Senegal [56] and China [57,58]. If focusing on specific geographic areas has the advantage of exploring within a range of genotypes well adapted to that environment, examining wider collections opens the possibility of investigating the genetic relationship across landraces spread around the world, and having a more precise estimation of the genetic diversity within the group of landraces and with respect to advanced breeding lines or modern cultivars. To quote some examples, studies carried out on panels of hundreds of landraces have been considered in durum and common wheat [35,59,60], barley [61,62] and rye [63]. In general, a higher genetic diversity has been observed in the group of landraces compared to the groups of advanced breeding lines and modern cultivars, indicating landraces as a useful source of variation for breeding. Additionally, when clustering and population structure analyses have been considered, the total genetic variation was higher within than between groups, and the groups were in general consistent with the geographical origin of the lines, except in a few cases. Mzid et al. [64] assessed genetic diversity in a panel of 53 Lebanese barley landraces through the electrophoretic pattern of the seed storage proteins, hordeins. In this case, the absence of correlation between the genetic variability and the geographic origin of sample provenance was explained by the fact that Lebanon is a small country where seeds are easily exchanged between farmers’ communities in the different regions. Similarly, Yadav et al. [47] phenotypically evaluated 25 naked barley landraces from different regions of Nepal. The UPGMA cluster analysis, carried out with qualitative phenotypic descriptors and quantitative traits, categorized the landraces in five clusters with no distinct regional grouping patterns. In this case, principal component analysis revealed the quantitative traits, such as grain yield, plant height and earliness, and qualitative traits, such as grain colour, lemma awn/hood and lemma awn barbs, to be the principal discriminatory characteristics of the Nepalese naked barley landrace collection.

Phenotypic evaluation of landraces is important to identify sources of useful loci for traits of interest in breeding and pre-breeding programs, in relation to traits with a simple genetic basis as the resistance to diseases, but also to complex traits such as grain yield (reviewed by Dwiwedi et al. [5]). Nevertheless, a clearer picture, in terms of genetic diversity, can be achieved using molecular markers. Markers based on polymorphisms at the level of seed storage proteins have been used in different cereal species such as Ethiopian emmer accessions [49], durum wheat and barley landraces [64,65]. Molecular markers based on DNA have been developed, both at the chromosome and the DNA sequence level. Polymorphisms were identified at the level of chromosome banding, through cytofluorometry [66,67]. The analysis of 58 varieties and landraces demonstrated a remarkable reproducibility and sensitivity of flow cytometry for the detection of numerical and structural chromosome changes [68]. In this regard, the dissection of complex genomes by flow cytometric sorting into the individual chromosomes reduces its complexity in a lossless manner, having a significant impact in many areas of research and giving a strong impulse to the sequencing of complex plant genomes [69,70,71]. At sequence level, DNA-based molecular markers have become the most suitable tool in this kind of study, thanks to their informativeness and to the great reduction in time processing and costs observed in the last few years. Random Amplified Polymorphic DNA (RAPD) markers were initially used for assessing genetic diversity in cereal landraces [49], but they were characterized by a low reproducibility, therefore Simple Sequence Repeat (SSR) markers became the method of choice thanks to their reproducibility and informativeness with a high number of alleles detected per locus. As an example, 8.1 alleles per locus were detected in a panel of 66 barley landraces from Tunisia [50], and 14.6 alleles per locus were identified in a collection including 36 oat commercial varieties and 141 landraces from Spain [53]. In more recent times, high-throughput methods have been developed, such as those based on fixed markers arrays, which include Diversity Array Technology (DArT) markers and SNP arrays. These methods have been shown to be suitable for genetic studies on cereal landraces and can assess a large number of entries, as in the case of panels with several hundreds of durum wheat landraces from Spain, assessed with DArT markers, or from Ethiopia and different countries worldwide, tested with SNP arrays [35,39]. An important aspect is that a certain ascertainment bias should be considered, as these platforms were mainly developed starting from cultivars [34]. For this reason, methods based on Genotyping by sequencing, including DArTseq, are also used in this kind of study [46,72]. The use of high-throughput markers allowed, in particular, the collection of more precise information on decay of linkage disequilibrium in landrace panels, which showed a higher resolution compared to commercial cultivars, for their use in association mapping analysis [51,73]. Moreover, the availability of a large number of markers with a good coverage of the genome is important to identify rare and private alleles, which are present only in a defined group of genotypes [35,36,37,38,39,40,41,42,43,44,45,46,47,48,49,50,51,52,53,54,55,56,57,58,59,60]. This kind of knowledge is very important for breeding, as landraces can be chosen not only based on their diversity per se, but also for specific alleles of interest in a particular breeding program.