PCR analysis of oilseed rape cultivars (Brassica napus L. ssp. oleifera) using 5?-anchored simple sequence repeat (SSR) primers

Charters, Y. M.; Robertson, A.; Wilkinson, M. J.; Ramsay, G.

doi:10.1007/s001220050147

Cited by 63 publications

(79 citation statements)

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“…ISSR-PCR analysis was performed according to the protocol described by Charters et al (1996) using one or two of the primers listed in Table 1. All possible pairwise combinations of primers were used on all three genotypes studied.…”

Section: Dna Extraction and Issr-pcr Analysismentioning

confidence: 99%

“…Amplification products were loaded onto a pre-cast polyacrylamide gel comprising a 5% w/v polyacrylamide stacking gel and a 10% w/v pre-cast polyacrylamide gel (Amersham Pharmacia Biotech, St Albans, U.K., Cleangel 48S). They were then fractionated and silver-stained according to the procedure described by Charters et al (1996). Two to four replicates were performed of each PCR on each genotype.…”

Section: Dna Extraction and Issr-pcr Analysismentioning

confidence: 99%

“…Mixes were made containing each primer in the following concentration ratios: 0: 0.2 µM; 0.1 µM: 0.2 µM; 0.2 µM: 0.2 µM; 0.2 µM: 0.3 µM; 0.2 µM: 0.4 µM. PCR master mixes were made using the protocol described by Charters et al (1996) but, where necessary, the final volume was adjusted to 20 µl by changing the amount of water used.…”

Section: Dna Extraction and Issr-pcr Analysismentioning

confidence: 99%

“…The cumulative number of extra markers produced by each primer when used in pairwise combinations is given at the base of the column for each primer Biotech, St Albans, U.K.) and silver nitrate-stained according to Charters et al (1996). Selected bands were then excised and reamplified by PCR using the same ISSR primers and PCR conditions described above.…”

Section: Dna Extraction and Issr-pcr Analysismentioning

confidence: 99%

“…The many amplification products generated by ISSR-PCR consist largely of the region between neighbouring and inverted SSR sites. Zietkiewicz et al (1994) first used this approach to distinguish between closely related plant genotypes but it has since been applied for cultivar identification (Wolff et al 1995;Kantety et al 1995;Charters et al 1996), to detect gene-flow and introgression (Timmons et al 1996;Allainguillume et al 1997) and for mapping purposes (Kojima et al 1998). The utility of this approach stems from its simplicity and reproducibility, the high number of polymorphisms revealed and the fact that there is no prior need for DNA sequence information from the organism under study.…”

Section: Introductionmentioning

confidence: 99%

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The potential of ISSR-PCR primer-pair combinations for genetic linkage analysis using the seasonal flowering locus in Fragaria as a model

Çekiç¹,

Battey²,

Wilkinson³

2001

Theor Appl Genet

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ISSR-PCR has been widely used for genetic distance analysis and DNA fingerprinting but has been less well utilised for mapping purposes. A key limitation lies in the small number of primer designs available to generate useful polymorphisms. In this study, the potential of paired combinations of ISSR primers is evaluated using a test cross mapping population of 168 BC 1 individuals between Fragaria vesca f. vesca and a closely related line F. vesca f. semperflorens. Ten ISSR primers and all possible pairwise combinations between them were used to generate markers potentially linked to the locus controlling seasonal flowering in F. vesca. Band profiles of individual primers were found to be highly reproducible for band position and intensity, and only minor variation was noted in band intensity (but not in position) between different constituent mixes of primer-pair combinations. Overall, ISSR primers used in isolation produced 85 markers of which only five were specific to F. vesca. None of these markers were linked to the seasonal flowering locus. In contrast, the primer-pair combinations yielded 493 markers, including 14 specific to F. vesca. These markers included two located within 2.2 cM of the seasonality locus. The strengths and limitations of using pairs of ISSR primers in combination for mapping and other genetic analyses are briefly explored.

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Section: Dna Extraction and Issr-pcr Analysismentioning

confidence: 99%

Section: Dna Extraction and Issr-pcr Analysismentioning

confidence: 99%

Section: Dna Extraction and Issr-pcr Analysismentioning

confidence: 99%

Section: Dna Extraction and Issr-pcr Analysismentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

The potential of ISSR-PCR primer-pair combinations for genetic linkage analysis using the seasonal flowering locus in Fragaria as a model

Çekiç¹,

Battey²,

Wilkinson³

2001

Theor Appl Genet

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The development of multiplex simple sequence repeat (SSR) markers to complement distinctness, uniformity and stability testing of rape (Brassica napus L.) varieties

et al. 2003

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To assess the potential of multiplex SSR markers for testing distinctness, uniformity and stability of rape (Brassica napus L.) varieties, we developed three multiplex SSR sets composed of five markers each. These were used to measure the extent of diversity within and between a set of ten varieties using a fluorescence-based semi-automated detection technology. Also, we evaluated the significance of any correlation between SSRs, pedigree and five of the morphological characters currently used for statutory distinctness, uniformity and stability testing of rape varieties. An assignment test was allowed to identify 99% of the plants examined, with the correct variety based on the analysis of 48 individual plants for each variety. Principal coordinate analysis confirmed that a high degree of separation between varieties could be achieved. Varieties were separated in three groups corresponding to winter, spring and forage types. These results suggested that it should be possible to select a set of markers for obtaining a suitable separation. Diversity within varieties varied considerably, according to the variety and the locus examined. No significant correlation was found between SSR and morphological data. However, genetic distances measured by SSRs were correlated to pedigree. These results suggested that SSRs could be used for pre-screening or grouping of existing and candidate varieties, allowing the number of varieties that need to be grown for comparison to be reduced. Multiplex SSR sets gave high-throughput reproducible results, thus reducing the costs of SSR assessment. Multiplex SSR sets are a promising way forward for complementing the current variety testing system in B. napus.

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The development of ISSR-derived SCAR markers around the SEASONAL FLOWERING LOCUS (SFL) in Fragaria vesca

2004

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Fragaria vesca is a short-lived perennial with a seasonal-flowering habit. Seasonality of flowering is widespread in the Rosaceae and is also found in the majority of temperate polycarpic perennials. Genetic analysis has shown that seasonal flowering is controlled by a single gene in F. vesca, the SEASONAL FLOWERING LOCUS ( SFL). Here, we report progress towards the marker-assisted selection and positional cloning of SFL, in which three ISSR markers linked to SFL were converted to locus-specific sequence-characterized amplified region (SCAR1-SCAR3) markers to allow large-scale screening of mapping progenies. We believe this is the first study describing the development of SCAR markers from ISSR profiles. The work also provides useful insight into the nature of polymorphisms generated by the ISSR marker system. Our results indicate that the ISSR polymorphisms originally detected were probably caused by point mutations in the positions targeted by primer anchors (causing differential PCR failure), by indels within the amplicon (leading to variation in amplicon size) and by internal sequence differences (leading to variation in DNA folding and so in band mobility). The cause of the original ISSR polymorphism was important in the selection of appropriate strategies for SCAR-marker development. The SCAR markers produced were mapped using a F. vesca f. vesca x F. vesca f. semperflorens testcross population. Marker SCAR2 was inseparable from the SFL, whereas SCAR1 mapped 3.0 cM to the north of the gene and SCAR3 1.7 cM to its south.

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PCR analysis of oilseed rape cultivars (Brassica napus L. ssp. oleifera) using 5?-anchored simple sequence repeat (SSR) primers

Cited by 63 publications

References 0 publications

The potential of ISSR-PCR primer-pair combinations for genetic linkage analysis using the seasonal flowering locus in Fragaria as a model

The potential of ISSR-PCR primer-pair combinations for genetic linkage analysis using the seasonal flowering locus in Fragaria as a model

The development of multiplex simple sequence repeat (SSR) markers to complement distinctness, uniformity and stability testing of rape (Brassica napus L.) varieties

The development of ISSR-derived SCAR markers around the SEASONAL FLOWERING LOCUS (SFL) in Fragaria vesca

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