Characterization of Microsatellite Markers and Their Application for the Assessment of Genetic Diversity among Lotus Accessions

To evaluate the genetic diversity and relatedness of 11 types of lotus (Nelumbo Adans.) accession, which represent the majority of lotus genotypes cultivated in China, 78 microsatellite-containing clones from a small insert library of Nelumbo nucifera Gaertn. enriched for AC/GT repeats were isolated and sequenced. Twenty-four primer pairs were designed and nine polymorphic simple sequence repeat (SSR) markers were amplified successfully from Nelumbo lutea Pers. and N. nucifera genomic DNA. These nine SSR markers and an additional two SSR markers obtained from GenBank showed a high degree of polymorphism. Forty-one alleles were detected, which yielded a mean of 3.7 alleles per locus. The polymorphic information content varied from 0.157 to 0.785 (mean, 0.537), which indicated a high level of informativeness. The 71 lotus accessions revealed a high level of genetic diversity, among which the Sino-American hybrids showed the highest genetic diversity (effective number of alleles = 2.760, percentage of polymorphic loci = 100%, expected heterozygosity = 0.610). The unweighted pair group method with arithmetic mean cluster and principal coordinate analyses both revealed that the wild lotus accessions were genetically distinct from the cultivated genotypes and probably suffered genetic intrusion from introduced cultivars. Thus, in situ conservation should be undertaken to protect the wild resources. In the cluster analysis, the three tropical lotus cultivars were separated from the wild genotypes of N. nucifera and all the genotypes of N. lutea, which indicated that the tropical ecotypes were genetically distinct from the temperate ecotypes and should be used as parents for the introduction of special traits by breeding. Two types of N. nucifera, versicolor lotus and thousand petals lotus, were clustered together. Giventhat theflower characteristics of these accessions were correlatedstrongly withtheir genetic constitution, they should be introduced extensively into breeding programs to create novel cultivars. The genus Nelumbo has received much attention for its special position in the early stages of angiosperm diversification (Hayes et al., 2000). Only two species are extant:N.nucifera and N.lutea. These species are relics of the ice age and are considered to be living fossils (Xue et al., 2006). The sacred asian lotus (N. nucifera) occurs in warm-temperate to tropical regions of Asia and northeast Australia. The american lotus (N. lutea )i s confined to eastern North America from southern Canada to Florida (Borsch and Barthlott, 1994). The american lotus is seldom cultivated as an ornamental plant and is considered to