Highly informative single-copy nuclear microsatellite DNA markers developed using an AFLP-SSR approach in black spruce (Picea mariana) and red spruce (P. rubens) (original) (raw)
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Molecular Ecology Notes, 2005
The spruce ( Picea ) species are ecologically and economically important in Canada. Highly informative markers with high multiplex ratios are needed to assist spruce genomics, genetics, and breeding programs. Selectively amplified microsatellite polymorphic loci (SAMPL) markers are highly suitable for these programs. We have developed, optimized, and characterized a set of 10 new SAMPL primers in combination with 16 Mse I primers and resolved a large number of polymorphic SAMPL markers in spruce. The SAMPL primers were designed from the compound microsatellite repeats found in Norway spruce ( Picea abies ) and white spruce ( Picea glauca ). A total of 6313 polymorphic SAMPL makers were produced by 160 SAMPL-Mse I primers combinations in eight progeny of a spruce mapping population.
Theoretical and Applied Genetics, 2002
Trinucleotide microsatellites have proven to be the markers of choice in human genetic analysis because they are easier to genotype than dinucleotides. Their development can be more time-consuming due to their lower abundance in the genome. We isolated trinucleotide microsatellites in Norway spruce (Picea abies K.) using an enrichment procedure for the genomiclibrary construction. Here we report on the characterisation of 85 ATC microsatellite-containing clones, from which 39 markers were developed. Many of the clones showed the occurrence of tandem repeats of higher order than the trinucleotide ones, often resembling minisatellite repeats. The sequencing of a sample of the alleles at one of the loci revealed size homoplasy due to base substitutions within the microsatellite region. The presence of ATC motifs within repetitive sequence families was observed. We found a significant relationship between the level of polymorphism and the length of the microsatellite. The levels of variability for ATC trinucleotide markers were lower than those for dinucleotides, both when tested on all loci in a set of six individuals and on a subset of loci in four natural populations. This difference is most likely attributable to lower mutation rates for trinucleotide than for dinucleotide loci. The availability of markers with different mutation rates allows one to select the proper marker set to investigate population processes on different time scales.
Tetranucleotide and Dinucleotide Microsatellite Markers for Red Spruce (Picea rubens
Three library enrichment protocols were used to develop tetra-and dinucleotide microsatellite markers for red spruce (Picea rubens). Two enrichment methods based on a single round of magnetic beads-based subtractive hybridization showed limited success. A third method with two successive rounds of subtractive hybridization had a high enrichment success but was found to increase the rate of redundancy and chimerical sequences among the clone sequences. Several single locus tetra-and dinucleotide markers were discovered and can be useful in mapping studies. All newly developed markers except one were characterized by a high frequency of null alleles. One new and one previously described microsatellite marker were identified as informative, reliable and characterized by a low null allele frequency, and were suitable for paternity/parentage and population genetic analyses in red spruce.
Theoretical and Applied Genetics, 2002
The development of microsatellite markers can be a time-consuming process, especially in species such as conifers where many microsatellites have been shown to be associated with the repetitive fraction of the genome and to produce complex banding patterns following electrophoresis. Therefore, procedures to eliminate this fraction from further processing are sought. In this paper, we report on the development of 53 dinucleotide SSR markers in Norway spruce, 35 of which (66%) produce simple, polymorphic patterns. This high efficiency is obtained by introducing a dot-blot selection against high copy number sequences, performed on the microsatellite-containing clones. The resulting markers turned out to be polymorphic and useful for population genetic studies and for linkage mapping. Seven additional markers that were not subject to the dot-blot selection are also presented.
Silvae Genetica
Fifteen previously published microsatellite primer pairs developed for and tested on Canadian white spruce were screened for amplification and polymorphy in Alaskan populations and tested for their suitability in PCR multiplexing. Eleven loci expressing polymorphisms ranging from 7 to 58 alleles were selected for development and optimization of three multiplex assays. Four natural stands containing a total of 1470 trees were used to characterize the selected loci and demonstrate their applicability for genotyping studies and parentage analysis. These assays can be used for studies focusing on population genetics, parentage analysis, provenance research, or individual genetic fingerprinting. The use of multiplex PCR facilitates large-scale studies by simultaneously enabling high resolution and reducing processing time and per sample cost.
Molecular Ecology Notes, 2004
Few microsatellite markers have been specifically developed for Picea sitchensis. In January 2004 the appearance of over 10 000 sequences of expressed regions of DNA from P. glauca in GenBank presented an opportunity for the development of additional microsatellite markers in Sitka spruce. Mono‐ and dinucleotide repeat sequences were located in these sequences and primers were designed around these regions. Amplification was attempted in Sitka material from a broad geographical range and the level of polymorphism was assessed. Primers were also tested in progeny of a controlled cross. Nine polymorphic loci that demonstrated Mendelian inheritance in Sitka were discovered in this study.