Summary
Pteris vittata was the first identified arsenic (As) hyperaccumulator. Here we investigated whether phytochelatins (PCs) are involved in the hypertolerance of arsenic by P. vittata.
P. vittata was exposed to 0–500 µm arsenate for 5 d, or to 50 µm arsenate for 0–7 d. In addition, l‐buthionine‐sulphoximine (BSO), an inhibitor of γ‐glutamylcysteine synthetase, was used in combination with different arsenate exposures. The relationships between As accumulation and the concentrations of PCs and glutathione (GSH) were examined.
PC synthesis was induced upon exposure to arsenate in P. vittata, with only PC2 detected in the plant. The As concentration correlated significantly with PC2 concentration in both roots and shoots, but not with GSH. The molar ratio of PC‐SH to As was c. 0.09 and 0.03 for shoots and roots, respectively, suggesting that only a small proportion (1–3%) of the As in P. vittata can be complexed with PCs. In the presence of arsenate, addition of BSO decreased PC2 concentrations in roots and shoots by 89–96% and 30–33%, respectively. BSO alone was found to inhibit root growth of P. vittata markedly.
The results suggest that PCs play a limited role in the hypertolerance of As in P. vittata.
Amplified fragment length polymorphism (AFLP) analysis of 26 trees of three Salix taxa: Salix alba L. (White Willow), S. fragilis L. (Crack Willow) and their hybrid S. x rubens Schrank, across an example of their habitat range in south‐west Germany, supported the distribution previously determined using morphological characterization. UPGMA and principal coordinates analysis of the AFLP data revealed three distinct clusters corresponding to the three taxa. In addition, AFLP analysis on individuals which were difficult to identify morphologically revealed that they were either the hybrid S. x rubens or S. fragilis. Four specimens of S. fragilis were indistinguishable with three primer combinations suggesting they are members of one clone.
Microsatellites or simple sequence repeats (SSRs) were isolated from coconut (Cocos nucifera) and tested for polymorphism on restricted germplasm. Sequencing of 197 clones from a cv. Tagnanan Tall-enriched genomic library showed that 75% contained a microsatellite, of which 64% were dinucleotide (GA/CT, CA/GT and GC/CG), 6% were trinucleotide, and 30% were compound repeats. Of 41 primer pairs tested on Tagnanan Tall genomic DNA, 38 gave the expected size product, two amplified two loci, and another gave a multilocus pattern. On 20 coconut samples, the 38 SSRs detected 198 alleles (average: 5.2 alleles per microsatellite). Genetic diversity (D = 1 - sigma pi2) values ranged from 0.141 to 0.809. Heterozygotes were present at high frequencies among some dwarf samples. Analysis of similarity matrices based either on shared alleles at each locus (simple matching coefficient) or on allele bands across all loci (Jaccard coefficient) showed similar results. Dwarfs grouped separately from talls and showed less genetic diversity. In a wider test on 40 samples, 8 SSRs detected 64 alleles (average: eight alleles per microsatellite). These results indicate the high potential of microsatellites to detect genetic diversity in coconut germplasm.
The genus Salix (willow) contains a number of species of great value as biomass crops. Efforts to breed varieties with improved biomass yields and resistances to pests and diseases are limited by the lack of knowledge on the genetic basis of the traits. We have used AFLP and microsatellite markers to construct a genetic map of willow from a full-sib cross of the diploid species Salix viminalis (2n = 38). In accordance with a double pseudo-testcross approach, separate parental maps were constructed and merged to produce a consensus map comprising 291 AFLP and 39 willow microsatellite markers. Nineteen poplar microsatellites were also tested in willow. Five of these amplified loci, of which two were mapped. Linkage groups of the consensus map that could be identified in the parental maps are presented here and spanned 1,256.5 cM with an average interval between markers of 4.4 cM.
Forty‐six microsatellites were isolated from an enriched library of Salix burjatica and tested on 20 individuals (of nine species/hybrids) from the National Willows Collection (IACR‐Long Ashton Research Station, UK). Twenty‐nine were monomorphic, gave multilocus or unscorable patterns, or were duplicates. The remaining 17 microsatellites gave 2–22 alleles/locus. Three microsatellites successfully cross‐amplified in 31 additional Salix species. A further six were tested on panels comprising 6–25 individuals from the 31 species. Cross‐amplification was successful in all cases. These results suggest that the microsatellites isolated here should prove useful for population studies in a wide range of Salix species.
The genus Salix (willow) contains a number of species which have great potential value as biomass crops in short rotation coppice (SRC). Efforts to improve biomass willows by breeding are currently hampered by the limited information available on genetic diversity and on genetic relationships within and among species, clones, and hybrids in the gene pool. Hybridisation occurs commonly in nature and the relatedness of many clones is unclear. Molecular markers were used to assess genetic diversity in a reference set of willows maintained within the U.K. National Collection and 16 elite clones currently being evaluated in field trials at several European sites. The two marker systems tested, RAPDs and AFLPs, were equally informative for revealing relationships within the reference set of clones. No differences were observed when alternative similarity coefficients were compared or when analysis was restricted to the use of polymorphic bands only. Good agreement with available knowledge of the clonal origins was obtained and one instance of duplicate clones was identified. AFLPs revealed more genetic diversity and discriminated between closely related clones. A difference in the relationships revealed was observed with one AFLP primer combination. RAPDs were more problematic, both in terms of reproducibility and scorability.
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