Abstract
BackgroundMicrosatellites or single sequence repeats (SSRs) are a powerful choice of marker in the study of Phytophthora population biology, epidemiology, ecology, genetics and evolution. A strategy was tested in which the publicly available unigene datasets extracted from genome sequences of P. infestans, P. sojae and P. ramorum were mined for candidate SSR markers that could be applied to a wide range of Phytophthora species.ResultsA first approach, aimed at the identification of polymorphic SSR loci common to many Phytophthora species, yielded 171 reliable sequences containing 211 SSRs. Microsatellites were identified from 16 target species representing the breadth of diversity across the genus. Repeat number ranged from 3 to 16 with most having seven repeats or less and four being the most commonly found. Trinucleotide repeats such as (AAG)n, (AGG)n and (AGC)n were the most common followed by pentanucleotide, tetranucleotide and dinucleotide repeats. A second approach was aimed at the identification of useful loci common to a restricted number of species more closely related to P. sojae (P. alni, P. cambivora, P. europaea and P. fragariae). This analysis yielded 10 trinucleotide and 2 tetranucleotide SSRs which were repeated 4, 5 or 6 times.ConclusionKey studies on inter- and intra-specific variation of selected microsatellites remain. Despite the screening of conserved gene coding regions, the sequence diversity between species was high and the identification of useful SSR loci applicable to anything other than the most closely related pairs of Phytophthora species was challenging. That said, many novel SSR loci for species other than the three 'source species' (P. infestans, P. sojae and P. ramorum) are reported, offering great potential for the investigation of Phytophthora populations. In addition to the presence of microsatellites, many of the amplified regions may represent useful molecular marker regions for other studies as they are highly variable and easily amplifiable from different Phytophthora species.
Highlights
Microsatellites or single sequence repeats (SSRs) are a powerful choice of marker in the study of Phytophthora population biology, epidemiology, ecology, genetics and evolution
Other approaches to study intraspecific variability among Phytophthora species including RAPD-PCR and AFLP have proved valuable within a particular study but comparing results from one laboratory to another has always proved challenging with such fingerprinting tools [10,11,12,13]
Analysis of sequences from P. infestans, P. sojae and P. ramorum genome projects and scanning for homologous simple sequence repeats (SSRs) Predicted gene datasets from P. infestans http:// www.broad.mit.edu/P. sojae and P. ramorum http:// genome.jgi-psf.org/ were scanned for the presence of microsatellites defined as short tandem repeat motifs (SSRs) of 2–6 bp
Summary
Microsatellites or single sequence repeats (SSRs) are a powerful choice of marker in the study of Phytophthora population biology, epidemiology, ecology, genetics and evolution. Accurate studies based on the analysis of mitochondrial and nuclear DNA have resulted in a consensus of the phylogenetic relationships within the genus with a grouping into 10 genetically related clades accepted [2,3,9]. These studies were based on genes commonly conserved within a species and unsuitable to characterize intraspecific variability. Major advantages SSRs include: (i) multiple SSR alleles may be detected at a single locus using a simple PCR-based screen, (ii) SSRs are evenly distributed across the genome, (iii) they are codominant, (iv) very small quantities of DNA are required for screening, (v) analysis may be semi-automated, and (vi) results are objective compared to random amplification methods [18]
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