Contributed by Sybil Herrera-Foessel, Ravi Singh and Evans Lagudah
Lr68 is an adult plant resistance (APR) conferring slow rusting resistance to wheat leaf rust caused by Puccinia triticina. This gene, formerly designated LrP, was first described in CIMMYT's spring bread wheat Parula (FKN/3/2*Frontana//KENYA 350 AD.9C.2/Gabo 55/4/Bluebird/Chanate). Parula is a line developed at CIMMYT in 1981 that combines also Lr34 and Lr46 (1-3). The likely origin of Lr68is the Brazilian cultivar Frontana (4)
Lillemo et al. (5) tested the effect of Lr68, Lr34 and Lr46 on leaf rust in an F6 recombinant inbred line population derived from a Avocet-YrA x Parula cross in nine field environments in Mexico, Brazil, Argentina, Uruguay and Chile. The authors were able to confirm additive effects of Lr68 with the other two slow rusting genes at each site, and also showed that in sites in Argentina and Uruguay, Lr68showed a stronger effect than Lr34.
Herrera-Fossel et al. (4) showed in field tests in Mexico that Lr68-carrying lines had lower severities to leaf rust infection with P. triticina races MCJ/SP and MBJ/SP than the susceptible checks. For most of the tests, the effect of Lr68 was smaller than those of Lr34, Lr46 and Lr67, and the combined effect of Lr34, Lr46 and Lr68 in Parula resulted in near immunity. These authors also demonstrated that Lr14b, a race-specific resistance gene, is a closely linked but different locus.
Markers for Lr68
Herrera-Fossel et al. (4) established the genetic linkage between Lr68 and nine markers. The closest loci flanking Lr68 are the CAPS marker Psy1-1 (0.5 cM proximal) and microsatellite gwm146 (0.6 cM distal). However, the PCR amplification for Psy1-1 does not yield consistent results and gwm146 is multi-allelic, so that finding the correct band associated with Lr68 is difficult. In consequence, the recommendation for MAS breeding are the co-dominant CAPS marker cs7BLNLRR or the dominant STS marker csGS, both proximal at 0.8 and 1.2 cM from Lr68 respectively.
Primers:
csGS-F1 5'- AAG ATT GTT CAC AGA TCC ATG TCA -3'
csGS-R1 5'- GAG TAT TCC GGC TCA AAA AGG -3'
cs7BLNLRR-F 5'- GAA GGA GTG CTT CCT CCA CTG -3'
cs7BLNLRR-R 5'- CTT GGT TCT CCT GTT CTT CCC -3'
PCR conditions:
- Denaturing step: 93°C, 1 min
- Amplification step (30 cycles):
- 93°C, 30 sec
- 60°C (csGS and cs7BLNLRR), 1 min
- 72°C, 1 min
- Extension step: 72°C, 5 min
The PCR product of the CAPS marker cs7BLNLRR was digested by adding a premix of 0.5 µl HaeIII restriction enzyme, 2 µl of 10X restriction buffer and 8 µl of water, and incubated at 37°C for 60 min on the PCR thermocycler.
PCR reaction mix:
- Taq Buffer 1X
- MgCl2 2.5 mM
- dNTP Mix 200 µM each
- Taq Enzyme 1U
- Glycerol (optional) 2.5%
- Primers F + R 0.25 µM each
- DNA 50 ng
Expected products
The PCR products were loaded on 12% acrylamide (29:1) gels, separated by electrophoresis and the bands were visualized using silver staining.
The dominant marker csGS yields a product with an expected size of about 385 bp in gentic backgrounds carrying Lr68. The CAPS marker cs7BLNLRR yields a fragment of 738 in lines with Lr68, and two bands of 478 and 270 bp in lines without Lr68.
References
1. Detection of quantitative trait loci associated with leaf rust resistance in bread wheat. William HM, Hoisington D, Singh RP, Gonzalez de Leon D. In: Genome, 1997, 40:253-260. DOI: 10.1139/g97-036
2.Characterization of genes for durable resistance to leaf rust and yellow rust in CIMMYT spring wheats. William HM, Singh RP, Huerta-Espino J, Rosewarne G. In: Buck HT, Nisi JE, Salomon N (eds), 2007, Developments in Plant Breeding Vol 12 Wheat production in stressed environments, pp 65-70, Springer, Dordrecht, The Netherlands
3.Characterization and mapping of a gene component for durable leaf rust resistance in chromosome arm 7BL. Herrera-Foessel SA, Singh RP, Huerta-Espino J, Lagudah ES. In: Phytopathology, 2009, 99:S53-S53. [link to meeting abstracts]
4. Lr68: a new gene conferring slow rusting resistance to leaf rust in wheat. Herrera-Foessel SA, Singh RP, Huerta-Espino J, Rosewarne GM, Periyannan SK, Viccar L, Calvo-Salazar V, Lan C, Lagudah ES. Theoretical and Applied Genetics, 2012, 124:1475-1486. DOI 10.1007/s00122-012-1802-1
5.Multiple rust resistance and gene additivity in wheat: lessons from multi-location case studies in the cultivars Parula and Saar. Lillemo M, Singh RP, William M, Herrera-Foessel SA, Huerta-Espino J, Germa´n S, Campos P, Chaves M, Madriaga R, Xia X,Liang S, Liu D, Li Z, Lagudah E. In: Global Rust Initiative Meeting, 2011, St. Paul, pp 111–120. [link to proceedings]