Molecular Markers Linked to Brown Stem Rot Resistance Genes, Rbs1 and Rbs2, in Soybean

Hanson et al., 1988), and PI 437970 (Willmot and Nickell, 1989), respectively. Brown stem rot resistance at Brown stem rot (BSR) of soybean [Glycine max (L.) Merr.] is these loci is conditioned by dominant alleles, and the caused by the fungal pathogen Phialophora gregata (Allington & D.W. Chamberlain) W. Gams and occurs in soybean production areas presence of a dominant allele at any one of these three around the world. Brown stem rot resistance genes Rbs1, Rbs2, and loci has been associated with a resistant reaction to Rbs3 have been identified in soybean germplasm and plant introducBSR (Hanson et al., 1988; Willmot and Nickell, 1989). tions through traditional genetic analyses. Resistance to BSR has been Resistance thus appears to provide an efficacious, ecoshown to reduce yield losses in soybean, but selection for this trait is nomical means of control of BSR. laborious and confounded by environmental variation. The objectives Selection for BSR resistance is hampered by a high of this study were to identify molecular markers linked to BSR resislevel of environmental variation in the field (Chambertance genes Rbs1 and Rbs2, and map these genes in the soybean lain and Bernard, 1968) and labor-intensive, time-congenome. Genetic families of populations segregating for Rbs1 and suming assays in the greenhouse (Sebastian and Nickell, Rbs2 were evaluated in the greenhouse for BSR phenotypic reaction 1985; Sebastian et al., 1985). However, the use of molecand identified as resistant, segregating, or susceptible. Leaf tissue collected from members of F2:3 families was bulked and DNA simple ular markers linked to Rbs genes could accelerate selecsequence repeat (SSR) marker analysis was used to identify markers tion and eliminate the effects of environmental variathat cosegregated with BSR reaction phenotypes. Five pairs of Rbs2 tion. Molecular markers could also facilitate pyramiding near-isogenic lines were subjected to a similar analysis to verify results of Rbs loci which could provide more temporally or obtained from marker analysis conducted on the population segregatgeographically stable resistance to P. gregata in the fuing for Rbs2. Results of marker analyses indicated that SSR markers ture. Stable BSR resistance sources are desirable given Satt215 and Satt431 were linked to Rbs1 and that Satt244 and Satt431 the historical failure of many monogenic disease resiswere linked to Rbs2. Marker-assisted selection in the Rbs1 (using tance mechanisms in plants and reports of physiological Satt431) and Rbs2 (using Satt244) populations would have correctly specialization in P. gregata (Gray, 1971; Willmot et al., predicted 88 and 82%, respectively, of the BSR reaction phenotypes. 1989). There is also evidence that while single Rbs resisThe Rbs1 and Rbs2 loci map to Molecular Linkage Group J and lie in a region known to contain Rbs3. This region also contains loci tance alleles usually confer high-level resistance, they conditioning resistance to taxonomically diverse fungal pathogens and do not provide immunity from disease; there are reports a locus affecting nodulation in response to a bacterial symbiont. of BSR symptom development on soybean lines containing Rbs resistance alleles (Bachman et al., 1997a; Hanson et al., 1988; Nelson et al., 1989). B stem rot of soybean, caused by the soilborne Several different molecular marker systems have fungus Phialophora gregata, is an economically imbeen used successfully in soybean, although these sysportant disease in the north central USA. Yield losses tems can be limited by expense, labor requirements, as high as 38% have been recorded in fields with develor a lack of repeatability if used for marker-assisted opment of severe BSR symptoms (Bachman et al., selection (Denny et al., 1996; Mudge et al., 1997). Re1997b; Dunleavy, 1966; Gray, 1972; Mengistu et al., striction fragment length polymorphism (RFLP) mark1986; Weber et al., 1966). Soybean yield loss to BSR in ers were among the first molecular markers to be used the USA in 1994 was 260 000 Mg (Wrather et al., 1997), in soybean. However, RFLP markers have several disand yield loss to BSR in 1996, 1997, and 1998 was estiadvantages for use in marker assisted selection, includmated at approximately 837 500, 653 300, and 369 500 ing relatively low levels of polymorphism (Keim et al., Mg, respectively (Wrather and Stienstra, 1999). Resis1989, 1992) and time-consuming protocols (Mudge et tance to BSR has been identified and utilized in cultivar al., 1997). SSR markers are DNA sequences consisting development and germplasm enhancement. Three loci of short tandem repeats of two to five nucleotides (core designated Rbs1, Rbs2, and Rbs3 were identified in germsequences) flanked by conserved DNA sequences. plasm line L78-4094 (Hanson et al., 1988), PI 437833 These markers can vary in length, depending on the number of core sequences positioned in tandem arrangement. Sequence length polymorphism can be asM.S. Bachman, Syngenta Seeds, Inc., 317-330th St., Stanton, MN sayed by amplification of these regions with the con55018; J.P. Tamulonis, Monsanto, 634 East Lincoln Way, Ames, IA 50010; C.D. Nickell, Dep. of Crop Sciences, Univ. of Illinois, 1102 S. served flanking sequences used as primer templates in Goodwin Ave., Urbana, IL 61801; A.F. Bent, College of Agricultural the polymerase chain reaction (PCR) (Akkaya et al., and Life Sciences, UW-Madison, 1630 Linden Dr., Madison, WI 53706. 1992). SSR markers have been widely utilized in soyResearch, supported in part by the Illinois Soybean Program Operating Board, was from a thesis by the senior author in partial fulfillAbbreviations: AFLP, amplified fragment length polymorphism; ment of the requirements for the Ph.D. degree at the Univ. of Illinois. BSR, brown stem rot; MLG, molecular linkage group; PCR, polymerReceived. 11 May 1999. *Corresponding author (mike.bachman@ ase chain reaction; PI, plant introduction; QTL, quantitative trait syngenta.com). locus/loci; RFLP, restriction fragment length polymorphism; SSR, simple sequence repeat. Published in Crop Sci. 41:527–535 (2001).