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Oat SSR Markers

Oat (Avena sativa) Microsatellite Markers

 

Microsatellite markers (simple sequence repeat (SSR)) have been widely available for the genetic analysis of agronomically important cereal crops such as wheat, barley, and rice, and have formed the backbone of genetic studies in these crops due to their high levels of polymorphism and ease of use.  Despite the benefits of microsatellite markers, few are available for use in oat genomics, and even fewer have been placed on genetic maps, a pre-requisite for their efficient association with agronomically important traits.  The cross-applicability to oat of SSR markers developed from wheat, barley, and sorghum has generally been found to be poor.  Though the levels of polymorphism are adequate (if not high), the polymorphisms are often dominant in nature resulting from primer-template mis-match, as opposed to the co-dominant, multi-allelic polymorphisms that are generated at true microsatellite loci and from which SSR markers gain their analytical power.

We have developed a series of SSR markers based on oat DNA sequences.  These markers are freely available and information such as PCR primer sequences, amplicon sizes, annealing temperatures, and marker quality indications are listed on this site.

Marker polymorphism information is provided for markers that have been tested against the parental lines of several oat-mapping populations such as Kanota/Ogle, Ogle/TAM O-301, Marion/Terra, Mn841801-1/Noble-2, Aslak/Matilda, and CDC SolFi/HiFi.  Several of the markers listed here have not been polymorphic in genotypes tested by us to date but mediate good quality amplifications of single loci that may be useful against other genotypes.

Some of the markers have been mapped in selected populations and these are indicated as such in then polymorpohism tables.  We continue to map these markers and information is updated regularily on these pages.

 

The microsatellite markers were based on either oat genomic DNA sequences (AM series), or on oat EST sequences (cAM series).

 

AM Series SSR Markers

These markers were developed in 1999/2000.  SSRs were identified in DNA sequences obtained from three oat genomic DNA libraries enriched for (AC/TG)n, (AG/TC)n, or (AAG/TTC)n repeats.  DNA for library construction was obtained from the oat cultivar "Ogle".

 

Relevant references;

Li, C.D., B.G. Rossnagel, and G.J. Scoles.  2000.  The development of oat microsattelite markers and their use in identifying relationships among Avena species and oat cultivars.  Theor. Appl. Genet. 101:1259-1268.

Li, C.D., B.G. Rossnagel, and G.J. Scoles.  2000.  Tracing the phylogeny of the hexaploid oat Avena sativa with satellite DNAs.  Crop Sci. 40:1755-1763. 

 

cAM Series SSR Markers

These markers target SSRs identified from an EST sequence database generated in cooperation with the Natural Products Genomics Resource (NAPGEN) initiative of PBI-NRC.  Approximately 15,000 ESTs were isolated from the cultivar CDC Dancer and sequenced (single pass) yielding approximately 7000 unigenes.  EST sequences have been submitted to NCBI, (GenBank accession numbers GO581282-GO598990; GO59949-GO599494).  The EST database was mined for SSR containing regions using the Web-based "SSR Primer Discovery" tool, and  primers flanking the SSRs were designed through the integrated Primer 3 program (set at default values).  The analysis identified 260 EST sequences (4.2%) that contained perfect and/or imperfect SSRs of n ³ 6 for di-, n ³ 5 for tri-, n ³ 4 for tetra, and n ³ 4 for penta-nucleotide repeat motifs.  Sixty-eight percent of all repeats were tri-nucleotide motifs, 18.5% were dinucleotide motifs, 13% were tetra-nucleotide motifs and penta-nucleotide motifs was limited to one.  Approximately one third of the SSR containing sequences were BLASTX annotated to genes of known function.

One hundred and eighty SSR primer pairs were chosen on the basis of repeat lengths ³ 20 nucleotides (irrespective of motif length) to be assayed for marker quality and polymorhism potential.

Only those primer pairs that mediated the robust amplification of few (preferably single) loci are included in our list.  Most of the markers are assumed to amplify true SSR containing regions though this has not been confirmed via re-sequencing of amplification products.  However, all identified polymorphisms are at least co-dominant in nature.

 

Assay Conditions;

                PCR reactions consisted of 20 mM Tris-HCl, 50 mM KCl, 2 mM MgCl, 400 uM dNTP, each primer @ 200 nM, 1 unit of Taq polymerase, and 100 ng template DNA in a volume of 25 uL.  Amplification cycles (35X) comprised of 94°C (45 sec.), 55°C - 63°C (45 sec.), 72°C (60 sec.).  

 

Relevant references;

Eckstein, P., A. Beattie, B. Rossnagel, and G. Scoles.  2008.  Genic microsatellite markers from expressed sequence tags (ESTs) of developing oat seed.  8th International Oat Conference, Minneapolis, MN, June 28 - July 2, 2008.