Megahed H. Ammar

Molecular Diversity Assessment Using Sequence Related Amplified Polymorphism (SRAP) Markers in Vicia faba L.

Sequence-related amplified polymorphism (SRAP) markers were used to assess the genetic diversity and relationship among 58 faba bean (Vicia faba L.) genotypes. Fourteen SRAP primer combinations amplified a total of 1036 differently sized well-resolved peaks (fragments), of which all were polymorphic with a 0.96 PIC value and discriminated all of the 58 faba bean genotypes. An average pairwise similarity of 21% was revealed among the genotypes ranging from 2% to 65%. At a similarity of 28%, UPGMA clustered the genotypes into three main groups comprising 78% of the genotypes. The local landraces and most of the Egyptian genotypes in addition to the Sudan genotypes were grouped in the first main cluster. The advanced breeding lines were scattered in the second and third main clusters with breeding lines from the ICARDA and genotypes introduced from Egypt. At a similarity of 47%, all the genotypes formed separated clusters with the exceptions of Hassawi 1 and Hassawi 2. Group analysis of the genotypes according to their geographic origin and type showed that the landraces were grouped according to their origin, while others were grouped according to their seed type. To our knowledge, this is the first application of SRAP markers for the assessment of genetic diversity in faba bean. Such information will be useful to determine optimal breeding strategies to allow continued progress in faba bean breeding.

Faba bean genomics: Current status and future prospects

Faba bean represents a crucial source of protein for food, especially for Mediterranean countries, and local demand for faba bean grains is increasing. The crop is also gaining increased attention as an elite candidate for conservation agriculture. However, the complexity of the faba genome has made progress in breeding programs and molecular studies relatively slow compared with other legume crops. Recent advances in plant genomics have made it feasible to understand complex genomes such as faba bean. With the increase of faba bean consumption in the Middle East region, there is an urgent need to develop elite faba genotypes suitable for arid and semi arid environments, with high yield potential and acceptable nutritional quality. This article highlights the recent advances in legume and faba genomics and its potential to contribute to the above mentioned goal. Emphasis is given on prospects on faba improvements strategies from the Middle East point of view.

Morphological and molecular genetic variability analyses of Saudi lucerne (Medicago sativa L.) landraces

Abstract. Collection and characterisation of genetic resources are required for the development of new cultivars. We analysed genetic diversity among 18 non-dormant lucerne (alfalfa, Medicago sativa L.) accessions including ten local ecotypes and eight introduced accessions at morphological and molecular levels using sequence-related amplified polymorphism (SRAP) primers. Standardised canonical discrimination functions for the investigated morphological traits showed that the first function (explaining 75% of total variability among accessions) was strongly influenced by leaflet shape, stipule shape, and the peduncle : petiole length ratio. The 16 SRAP primer pair combinations generated 677 differently sized SRAP fragments (peaks), of which 665 (98.3%) were polymorphic across all 18 accessions. We detected high levels of polymorphism (average polymorphic information content value = 0.96, average of 42.3 polymorphic fragments per primer pair). Based on morphological and SRAP data, local accessions tended to group together in the same cluster or formed individual clusters. Clusters of local accessions at high similarity sometimes correlated with their collection site (Qasemi-2 and Qasemi-3) on molecular analysis. Results of cluster analysis based on SRAP showed no significant correlation with morphological characters based on the Mantel test (r = 0.04).This inconsistent clustering of accessions could be due to the allelic variation (presumably) in a small number of genes (24 traits) contributing to morphological characterisation, while the 677 SRAP fragments (loci) are assumed to be relatively widely distributed across the genome. The wide geographical distribution of lucerne populations across different environments may provide good genetic resources for breeding purposes. SRAP analysis was effective to study genetic variability of non-dormant lucerne. This information will be helpful in assessing selections for lucerne breeding programs to develop new cultivars adapted to harsh environmental conditions.

Phenological, Nutritional and Molecular Diversity Assessment among 35 Introduced Lentil (Lens culinaris Medik.) Genotypes Grown in Saudi Arabia

Morphological, nutritional and molecular analyses were carried out to assess genetic diversity among 35 introduced lentil genotypes (Lens culinaris Medik.). The genotypes exhibited significant differences for their field parameters and some of them showed noticeable superiority. The nutritional and proximate analysis showed that some genotypes were excellent sources of proteins, essential amino acids, minerals, anti-oxidants, total phenolic contents (TPC) and total flavonoid contents (TFC) and hence, highlights lentil nutritional and medicinal potential. Sequence-related amplified polymorphism (SRAP) and amplified fragments length polymorphism (AFLP) markers were used to estimate the genetic variability at the molecular level. The existence of a considerable amount of genetic diversity among the tested lentil genotypes was also proven at the molecular level. A total of 2894 polymorphic SRAP and 1625 AFLP loci were successfully amplified using six SRAP and four AFLP primer pair combinations. Polymorphism information content (PIC) values for SRAP and AFLP markers were higher than 0.8, indicating the power and higher resolution of those marker systems in detecting molecular diversity. UPGMA (unweighted pair group method with arithmetic average) cluster analysis based on molecular data revealed large number of sub clusters among genotypes, indicating high diversity levels. The data presented here showed that FLIP2009-64L and FLIP2009-69L could be used as a significant source of yield, total protein, essential amino acids, and antioxidant properties. The results suggest potential lentil cultivation in the central region of Saudi Arabia for its nutritional and medicinal properties, as well as sustainable soil fertility crop.

Inter-simple sequence repeat (ISSR)-based diversity assessment among faba bean genotypes

Thirty-four faba bean (Vicia faba L.) including local and exotic materials were subjected to molecular diversity assessment using 12 inter-simple sequence repeat primers. The molecular data showed unambiguous and qualitative (present or absent) fragments that gave repeatable patterns were considered for the analysis. The 12 selected primers produced a total of 71 fragments (loci), all of which were polymorphic using the 34 collected faba genotypes. The results of clustering Nei’s genetic distance using the unweighted pair group method with arithmetic average algorithm at the 0.52 dissimilarity separated genotypes to six main clusters with many subclusters. The local genotypes were distributed to most of all clusters. Genotypes collected from Egypt and King Saud University was grouped together in two clusters, ICARDA’s genotypes in two clusters and two genotypes (H8, local determent genotype and 987–255–95 line) formed a single cluster. The high number of subclusters formed in this study indicated that there is a high genetic variability related to collection sites and it should be utilised in faba bean improvement.

Assessment of genetic diversity among faba bean genotypes using agro-morphological and molecular markers

Forty faba bean (Vicia faba L.) genotypes were evaluated for their agro-morphological performance and molecular diversity under Central Region of Saudi Arabia conditions during 2010–11 and 2011–12 seasons. Field performance results showed that faba genotypes exhibited a significant amount of variation for their agro-morphological studied parameters. Giza40 recorded the tallest genotype (139.5 cm), highest number of seeds per plants (100.8), and the highest seed yield per plant (70.8 g). The best performing genotypes were Giza40, FLIP03-014FB, Gazira1 and Goff1. Genetic variability among genotypes was determined using Sequence Related Amplified Polymorphism (SRAP) and Amplified Fragment Length Polymorphism (AFLP) markers. A total of 183 amplified fragments (alleles) and 1758 polymorphic fragments (bands) in SRAP and 202 alleles and 716 bands in AFLP were obtained using six SRAP and four AFLP primer combinations respectively. Polymorphism information content (PIC) values for AFLP and SRAP markers were higher than 0.8, indicating the existence of a considerable amount of genetic diversity among faba tested genotypes. The UPGMA based clustering of faba genotypes was largely based on origin and/or genetic background. Result of cluster analysis based on SRAP showed weak and not significant correlation while, it was highly significant based on AFLP analysis with agro-morphological characters (r = 0.01, p > 0.54 and r = 0.26, p < 0.004 respectively). Combined SRAP and AFLP markers proved to be significantly useful for genetic diversity assessment at molecular level. They exhibited high discrimination power, and were able to distinguish the faba bean genotypes with high efficiency and accuracy levels.

In vitro rooting of leguminous plants: Difficulties, alternatives, and strategies for improvement

Leguminous plants include many of the most important food crops and are also cultivated for forage, used as a rich fiber source, and are important in the manufacture of pharmaceutical compounds. Their ability to biologically fix atmospheric nitrogen enhances soil fertility. Beyond conventional propagation methods, modern in vitro technology offers new means for propagating these plants. Significant progress has been achieved in somatic embryogenesis and embryo rescue, despite legumes’ notorious recalcitrance to transformation and regeneration of legumes. The establishment of stable and reproducible regeneration protocols has resulted in commercially successful propagation of some legume species. Still, regeneration protocols for many other legumes have not been successful, primarily due to poor in vitro rooting during regeneration. This review addresses the factors constraining rooting, describes alternative methods to promote in vitro rooting, and provides a summary of in vitro rooting studies performed across a large number of leguminous species.

Analysis of Expressed Sequence Tags (EST) in Date Palm

Expressed sequence tags (EST) were generated from a normalized cDNA library of the date palm Sukkari cv. to understand the high-quality and better field performance of this well-known commercial cultivar. A total of 6943 high-quality ESTs were generated, out of them 6671 are submitted to the GenBank dbEST (LIBEST_028537). The generated ESTs were assembled into 6362 unigenes, consisting of 494 (14.4%) contigs and 5868 (84.53%) singletons. The functional annotation shows that the majority of the ESTs are associated with binding (44%), catalytic (40%), transporter (5%), and structural molecular (5%) activities. The blastx results show that 73% of unigenes are significantly similar to known plant genes and 27% are novel. The latter could be of particular interest in date palm genetic studies. Further analysis shows that some ESTs are categorized as stress/defense- and fruit development-related genes. These newly generated ESTs could significantly enhance date palm EST databases in the public domain and are available to scientists and researchers across the globe. This knowledge will facilitate the discovery of candidate genes that govern important developmental and agronomical traits in date palm. It will provide important resources for developing genetic tools, comparative genomics, and genome evolution among date palm cultivars.

Development of Genomic Simple Sequence Repeats (SSR) by Enrichment Libraries in Date Palm

Development of highly informative markers such as simple sequence repeats (SSR) for cultivar identification and germplasm characterization and management is essential for date palms genetic studies. The present study documents the development of SSR markers and assesses genetic relationships of commonly grown date palm (Phoenix dactylifera L.) cultivars in different geographical regions of Saudi Arabia. A total of 93 novel simple sequence repeat (SSR) markers were screened for their ability to detect polymorphism in date palm. Around 71% of genomic SSRs are dinucleotide, 25% trinucleotide, 3% tetranucleotide, and 1% pentanucleotide motives and show 100% polymorphism. The Unweighted Pair Group Method with Arithmetic Mean (UPGMA) cluster analysis illustrates that cultivars trend to group according to their class of maturity, region of cultivation, and fruit color. Analysis of molecular variations (AMOVA) reveals genetic variation among and within cultivars of 27% and 73%, respectively, according to the geographical distribution of the cultivars. Developed microsatellite markers are of additional value to date palm characterization, tools which can be used by researchers in population genetics, cultivar identification, as well as genetic resource exploration and management. The cultivars tested exhibited a significant amount of genetic diversity and could be suitable for successful breeding programs. Genomic sequences generated from this study are available at the National Center for Biotechnology Information (NCBI), Sequence Read Archive (Accession numbers. LIBGSS_039019).