The Sr22 gene was originally identified in the diploid wheat species Triticum monococcum ssp. boeoticum accession G-21 (1) and T. monococcum L. accession RL5244 (2). It was then transferred to tetraploid and hexaploid wheat through interspecific hybridizations. Sr22 is effective against Ug99. However, a yield penalty and a delay in heading date associated with the T. monococcum ssp. boeoticum chromosome segment carrying Sr22 has limited its use (3, 8). Lines with Sr22 and reduced T. monococcum segments have been recently produced (4).
Sr22 was previously mapped on the long arm of chromosome 7A (5). Three linked markers, CFA2019, CFA2123 and BARC121 (6), were used for haplotyping this locus by Yu et al. (7), CFA2123 was not always polymorphic betwenn T. monococcum and Triticum aestivum. Olson et al. (4) produced a new set of lines with reduced alien fragments and found that the closest markers flanking Sr22 in these lines are WMC633 and CFA2123.
WMC633-F 5'- ACA CCA GCG GGG ATA TTT GTT AC -3 WMC633-R 5'- GTG CAC AAG ACA TGA GGT GGA TT -3CFA2123
CFA2123-F 5'- CGG TCT TTG TTT GCT CTA AAC C -3' CFA2123-R 5'- ACC GGC CAT CTA TGA TGA AG -3'CFA2019
CFA2019-F 5'- GAC GAG CTA ACT GCA GAC CC -3 CFA2019-R 5'- CTC AAT CCT GAT GCG GAG AT -3BARC121
BARC121-F 5'- ACT GAT CAG CAA TGT CAA CTG AA -3 BARC121-R 5'- CCG GTG TCT TTC CTA ACG CTA TG -3PCR conditions for CFA2019:
For a 12 µl PCR reaction:
Olson et al. (4) recently mapped Sr22 4 cM distal to CFA2123 and 1-2 cM distal to WMC633. Marker WMC633 produced a 117-bp allele in the source line Sr22Tb. This allele has not been observed in any T. aestivum samples evaluated thus far (Brown-Guedira, unpublished). Several alleles are amplified by WMC633 in wheat, ranging in size from 170 bp to 260 bp. Olson et al. (4) produced an Sr22 carrying line (U5616-20-154) where recombination occurred between the resistance gene and all the SSR markers mapped in the region. In the line U5616-20-154, with the smallest T. monococcum segment, SSR marker WMC633 shows a 229-bp fragment from the hard winter wheat parent Lakin that can be used as a marker to introgress Sr22 from this line.
|Amplification of marker WMC633 on wheat. Lane 1, 50 bp ladder; Lane 2, 117 bp resistant allele from Sr22Tb; Lanes 3-5, susceptible wheat lines with 171 bp, 193 bp, and 221 bp alleles, respectively; Lanes 6-8, wheat lines heterozygous for Sr22. Products were separated on a 1.5% agarose gel.|
PCR amplification by CFA2019 and CFA2123 showed 238 and 234 bp fragments, respectively, in the Sr22 source germplasm, Sr22TB. BARC121 amplified two polymorphic bands of 217 and 234 bp in most germplasm. However, only the 195-bp fragment was amplified in Sr22TB. Therefore, the haplotype for Sr22 was 238-234-195 for markers CFA2019, CFA2123 and BARC121, respectively (4). Since these markers are not completely diagnostic and may produce false positive result, care must be taken when interpreting the results. Also, the allele sizes reported in Ref. 7 are slightly different.
|Amplification of markers CFA2019 and BARC121. The allele sizes correspond to those reported in Ref. 7.|
Conditions presented here should be considered only as a starting point of the PCR optimization for individual laboratories.Distribution of Donor Germplasm
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