Original version contributed by Colin W. Hiebert
Markers for Lr67 - Yr46 - Sr55 - Pm46 - Ltn3
The Lr67 gene for adult plant resistance (APR) to leaf rust was identified in the common wheat accession PI250413 (1) and transferred into Thatcher to produce the backcross line RL6077 (Thatcher*6/PI250413). Lr67 is phenotypically similar to Lr34 because it could also be associated with resistance to stem rust (2) and stripe rust (3), although Lr67 confers a lower level of leaf rust resistance than that induced by Lr34 (4).
It was previously hypothesized that the gene in RL6077 could be Lr34 translocated from chromosome 7D to a different location, but later Lagudah et al. (5) showed that Lr34 is not present in RL6077. More recently Hiebert et al. (4) using a combination of molecular markers and cytogenetic analyses demonstrated that chromosome 7D is not involved in any translocation carried by RL6077. These authors mapped Lr67 on chromosome 4D. Their results also suggest that if the observed translocation breakpoints derive from PI250413, they could be linked to Lr67.
Hiebert et al. (4) used PCR assays of SSR loci on 4DS and 4DL ditelosomic lines complemented with deletion bin mapping to conclude that the size of the 4D introgression from PI250413 after five backcrosses into Thatcher includes part of the short arm and at least 40% of the long arm of chromosome 4D. They also showed that Lr67 is closest to Xcfd71 and probably on the distal side on chromosome arm 4DL, although further genetic mapping is required.
This map location is consistent with results obtained for the stripe rust resistance gene which was tightly linked to Xcfd71 in the H1777 population – an F4 line derived from a Thatcher x RL6077 cross–. Given that line H1777 also segregated for leaf rust resistance and carried a reduced donor segment, it is possible that both leaf and stripe rust resistance map to the same locus, as is the case withLr34-Yr18-Pm38 and Lr46-Yr29. Interestingly leaf tip necrosis, which is associated with Lr34 and Lr46, was also recorded in segregants carrying Lr67. Combinations of Lr34, Lr46, and Lr67 represent an attractive option to breeders for durable multi-pathogen resistance to leaf rust, stripe rust, stem rust, and powdery mildew.
Comparison of RL6077 with Thatcher and RL6106 (Thatcher + Lr34) over 4 years of field testing showed that both Lr34 and Lr67 conditioned improved resistance compared with Thatcher, with Lr34 conferring a higher level of resistance compared with Lr67. Also, field trials showed no significant effect of the APR gene on average yield and other agronomic traits, such as height, maturity, lodging, bulk density and kernel weight. Similarly, end-use quality traits (whole wheat, particle size and SDS sedimentation) did not differ significantly for both lines.
Herrera-Foessel et al (6) analized 74 RILs from a cross Avocet/RL6077 lacking Sr26 and found that the Lr67 locus also encodes for stem rust tolerance in adult plants, which was responsible for a 41% reduction of disease severity in a field test in Ciudad Obregon (Mexico) and a lower but still significant reduction in Njoro (Kenya) of 16% in the presence of race TTKST. In addition in other field experiments in Norway they demonstrated that this same locus caused a 20% average reduction in powdery mildew severity. Additionally this locus was associated with leaf tip necrosis. At this point it is not clear whether it is one pleiotropic gene or several tightly linked genes, They were designated Sr55, Pm46 and Ltn3.
Methods
A genome-wide scan of 372 SSR loci showed that Xcfd71-4D was significantly associated with Lr67 (p < 0.001). Further testing of adjacent markers reported on 4D in different recombinant populations and a screening of 247 wheat lines from diverse origins indicated that Xcfd71-4D and Xcfd23-4D are good choices as molecular markers for breeding (4).
Primers:
CFD71-F 5'- CAA TAA GTA GGC CGG GAC AA -3'
CFD71-R 5'- TGT GCC AGT TGA GTT TGC TC -3'
CFD23-F 5'- TAG CAG TAG CAG CAG CAG GA -3'
CFD23-R 5'- GCA AGG AAG AGT GTT CAG CC -3'
PCR conditions:
- Denaturing step: 94°C, 2 min
- Amplification step (30 cycles):
- 95°C, 1 min
- 60°C (CDF23 and CDF71), 60 sec
- 73°C, 1 min
- Extension step: 73°C, 5 min
PCR reaction mix:
- 0.8 mM each dNTP
- 1.5 mM MgCl2
- 50 mM KCl
- 2 pmol each primer
- 0.5 U Taq DNA polymerase
- 24 ng genomic DNA
Expected products
PCR products were separated by electrophoresis in 0.4 mm × 50 cM, 4% polyacrylamide (1×TBE) gels. PCR amplification of CFD71 in RL6077 yields a 214-bp allele from chromosome 4DL, and CFD23 amplifies a 211-bp fragment. The RL6077 allele of Xcfd71 was present in 22 lines of the panel of 247 wheat lines mentioned above, while the Xcfd23 allele was present in 13 lines. Interestingly, these 13 lines also carried the RL6077-like allele for Xcfd71-4D, suggesting that this marker haplotype was retained in breeding programs across the world. However, no conclusive genetic data are available to confirm if this represents the transmission of Lr67.
References
1. Adult-plant leaf rust resistance in PI 250413, an introduction of common wheat. Dyck PL, Samborski DJ. In: Canadian Journal of Plant Science, 1979, 59:329–332
2. An interchromosomal reciprocal translocation in wheat involving wheat leaf rust resistance gene Lr34. Dyck PL, Kerber ER, Aung T. In: Genome, 1994, 37:556–559. DOI:10.1139/g94-079
3. Genetic association of leaf rust resistance gene Lr34 with adult-plant resistance to stripe rust in bread wheat. Singh RP. In: Phytopathology, 1992, 82:835–838. [Journal link]
4. An introgression on wheat chromosome 4DL in RL6077 (Thatcher*6/PI 250413) confers adult plant resistance to stripe rust and leaf rust (Lr67). Hiebert CW, Thomas JB, McCallum BD, Humphreys DG, DePauw RM, Hayden MJ, Mago R, Schnippenkoetter W, Spielmeyer W. In: TAG Theoretical and Applied Genetics, 2010, 121:1083–1091. DOI:10.1007/s00122-010-1373-y
5. Gene-specific markers for the wheat gene Lr34/Yr18/Pm38 which confers resistance to multiple fungal pathogens. Lagudah ES, Krattinger SG, Herrera-Foessel S, Singh RP, Huerta-Espino J, Spielmeyer W, Brown-Guedira G, Selter LL, Keller B. In:TAG Theoretical and Applied Genetics, 2009, 119:889–898. DOI:10.1007/s00122-009-1097-z
6. Lr67/Yr46 confers adult plant resistance to stem rust and powdery mildew in wheat. Herrera-Foessel SA, Singh RP, Lillemo M, Huerta-Espino J, Bhavani S, Singh S, Lan C, Calvo-Salazar, V, Lagudah, ES. In: Theoretical and Applied Genetics, 2014, 127:781-789. DOI:10.1007/s00122-013-2256-9.