'PI 181434' is a spring wheat germplasm originating from Afghanistan that was resistant to of stripe rust in a series of screenings carried out in Washington from 2004 to 2009. The type of resistance was all-stage, and it was effective to races accounting for all types of virulence in that period.
Li et al. (1) crossed 'PI 181434' and the 'Avocet Susceptible' cultivar to produce a F2:3 population to map the resistance gene. In this population stripe rust resistance was controlled by a single dominant gene on chromosome arm 3DL designated Yr45.
In the pathogenicity tests Yr45 showed resistance to seven races of Puccinia striiformis f. sp. tritici. Among them was PST-100, which has been the most predominant race in the USA since 2005 (2) and PST-127, a race with virulences to 17 of the 20 differential genotypes.
A set of molecular markers including SSRs and RGAPs (resistance gene analog polymorphism) were scored. The closest markers to Yr45 were two RGAPS, Xwgp118 on the distal side and Xwgp115 on the proximal side, that mapped at 4.8 and 5.8 cM, respectively (see Figure). Both markers are dominant, and amplified PCR products in the 'PI 181434' parent. However, some lines that do not carry Yr45 also had one or both markers present. An analysis of the sequences of the amplification products from these lines and 'PI 181434' revealed the presence of some SNPs that could be used in these cases (see Figure 3 of reference 1). Also, in a paper published later that reported a genome wide association mapping for stripe rust resistance in a set of hexaploid wheats (3), the authors found another SNP (IWA3012, wsnp_Ex_c25945_35206573, 9k Illumina Infinium SNP assay) significantly associated to stripe rust resistance in a location similar to that of Yr45. Although it was not further studied or validated, this SNP could be considered as an alternative marker.
Since Xwgp115 and Xwgp118 are both dominant markers, not very closely linked to Yr45 and there are lines in which the gene is not present but still yield positive results for one or both markers, users are advised to carefully test their parents before incorporating these markers in a breeding program.
Markers for Yr45
Primer sequences:
Xwgp115
Pto-kin4 5'- AGT GTC TTG TAG GGT ATC -3'
NLRR-IN2 5'- TCA GGC CGT GAA AAA TAT -3'
Xwgp118
Pto-kin1IN 5'- AAG TGG AAC AAG GTT ACG -3'
RLRR Rev 5'- ACA CTG GTC CAT GAG GTT -3'
Final concentrations of the PCR mix
- 30 ng template DNA
- 3.0 µl Mg-free 5× PCR buffer (Promega Inc.)
- 0.6 unit of Taq DNA polymerase (Promega)
- 5 mM MgCl2 (Promega Inc.)
- 10.25 mM of each dNTP (Sigma Chemical Co., St. Louis, MO, USA)
- 30 ng of each primer synthesized (Operon Biotechnologies, Inc., Huntsville, AL, USA)
Reaction volume: 15µl
PCR conditions
- Denaturing step: 94°C, 5 min
- Amplification step (45 cycles):
- 94°C, 1 min
- 45°C (wgp115 and wgp118), 1 min
- 72°C, 2 min
- Extension step: 72°C, 7 min
Expected products
PCR reactions were performed in a GeneAmp® PCR System 9700 thermo-cycler. The PCR products were separated on a 5% polyacrylamide gel and silver-stained for visualization
When linked to Yr45, Xwgp118 amplifies a band of 411-bp, while Xwgp115 yields a band of 492-bp.
References
1. Yr45, a new wheat gene for stripe rust resistance on the long arm of chromosome 3D. Li Q, Chen XM, Wang MN, Jing JX. In: Theoretical and Applied Genetics, 2011, 122:189-197. DOI: 10.1007/s00122-010-1435-1.
2. Challenges and solutions for stripe rust control in the United States. Chen XM. In: Australian Journal of Agricultural Research, 2007, 58:648–655. DOI: 10.1071/AR07045.
3. Genome-wide association mapping for seedling and adult plant resistance to stripe rust in synthetic hexaploid wheat.Zegeye H, Rasheed A, Makdis F, Badebo A, Ogbonnaya FC. In: PLoS ONE, 2014, 9(8): e105593. DOI: 10.1371/journal.pone.0105593.