Analysis Of Genetic Diversity And Development Of Molecular Markers On Faba Bean (Vicia Faba L.) Germplasm

Faba bean (Vicia faba L.) is an important grain legume crop with high seed-protien content, and it cultivated in China for 2100 years. China is the largest producer in both sowing area and production of faba bean globally. Faba bean becomes an important cash crop in western provinces and underdeveloped mountainous areas of China, due to its effective nitrogen fixation, soil improvement abilities and environmental protection in modern cropping systems. Various kind of DNA markers have been successfully carried out in other legume crops such as soybean, common bean and pea, etc. However, the marker assisted genetic studies on faba bean is quite limited. The main goal of this study was to compare genetic relationship of winter ecotype faba bean accessions with spring ecotype faba bean germplasm from China, and to analyze the genetic diversity of global faba bean germplasm as whole, to evaluate their genetic relationship and population structure, based on ISSR markers, in order to providing essential information for germplasm evaluation and effective utilization of faba bean genetic resources. Although there were approximately 100 microsatellite markers in faba bean have been reported, the number of molecular markers currently available publicly for faba bean is insufficient for widespread application in genetic studies. Therefore, developing more reliable molecular markers is necessary. DNA fragments from ISSR amplification were sequenced. Based on these sequences data, STS primer pairs were designed. The polymorphism of the primer was identified in 32 faba bean accessions from China and abroad. The main results in this study were summarized as follows:1. Eleven ISSR primers generated 271 unambiguous bands in 802 faba bean landraces and varieties from different geographical locations of China and abroad, of which 253 were polymorphic, and the rate of polymorphic bands was 0.93. The genetic diversity parameters of faba bean accessions from different geographical groups indicated that accessions from north China showed the highest genetic diversity, while accessions from central China and Afcica showed low level of diversity. The genetic diversity parameters of faba bean accessions from different provinces of China were compared, the result demonstrated that accessions from Yunnan and Inner Mongolia showed a higher genetic diversity.2. Germplasm from China is quite distinct to that from exotic accessions, based on principal component analysis and UPGMA clustering analysis. Chinese spring faba bean germplasm was clearly separated from Chinese winter faba bean, winter accessions from Zhejiang and Sichuan were quite distinct to that from other provinces of China. The result indicated that the genetic relationship of faba bean in China were closely associated with their geographical origins. Accessions from the same province or neighbouring provinces mostly clustered together.3. The results of AMOVA analyses indicated that the majority of the genetic variation in faba bean was due to within population variation. There was a highly significant difference between Chinese populations and other populations from the rest of the world. Chinese winter ecotype populations showed the highest genetic differentiation among populations. The largest variation within populations was detected in spring and winter ecotype accessions from China. Chinese populations based on different province and growth habit showed a high level of genetic variation within populations than other populations from the rest of the world.4. The result of clustering analysis indicated that the genetic similarity and diversity of faba bean germplasm are closely associated with their growth habit, their geographical origin and ecological distribution. The germplasm resources from Asia, Europe, Africa and China are closely related to their geographical distribution. As a continental geographic group, accessions from Europe were genetically closer to those from North Africa. There is broad agreement with previous AFLP studies of faba bean cultivars in Asia, Europe (Northern and Southern) and North African origin.5. Unambiguous DNA fragments from ISSR amplification were excised and extracted, the purified fragments were then ligated into the PGEM-T plasmid vector and transformed into Escherichia coli TOP10 competent cells. Positive colonies were identified and selected. Finally, purified plasmid DNAs were sequenced. Based on these sequences data, STS primer pairs were designed by using primer5.0 software. A tolal of 20 novel STS loci was identified as polymorphism. The polymorphism of the primer was identified in 32 faba bean accessions from China and abroad.