P. Balint-Kurti et al Precise mapping of SLB QTL in maize p1 Precise mapping of Quantitative Trait Loci for Resistance to Southern Leaf Blight, caused by Cochliobolus heterostrophus race O, and Flowering Time using Advanced Intercross Maize Lines

The IBM population, an advanced intercross recombinant inbred line population derived from a cross between the maize lines B73 (susceptible) and Mo17 (resistant), was evaluated in four environments for resistance to southern leaf blight (SLB) disease caused by Cochliobolus heterostrophus race O. Two environments were artificially inoculated, while two were not inoculated and consequently had substantially lower disease pressure. Four common SLB resistance QTL were identified in all environments; two in bin 3.04 and one each in bins 1.10 and 8.02/3. There was no significant correlation between disease resistance and days to anthesis. A direct comparison was made between SLB QTL detected in two populations, independently derived from the same parental cross; the IBM advanced intercross population and a conventional recombinant inbred line population. Several QTL for SLB resistance were detected in both populations, with the IBM providing between five and, in one case, fifty times greater mapping resolution. P. Balint-Kurti et al Precise mapping of SLB QTL in maize p4 INTRODUCTION Quantitative trait locus (QTL) mapping using biparentally-derived populations has historically been imprecise, with the support or confidence interval for a QTL position spanning 10-30 cM or comprising 1-3% of a genome (KEARSEY and FARQUHAR 1998; DEKKERS and HOSPITAL 2002; SALVI and TUBEROSA 2005). Reasons for this level of imprecision include insufficient marker density and limited opportunities for recombination between closely linked loci due to the relatively small size of many mapping populations (often 100-200 individuals). Increasing QTL resolution while maintaining a manageable population size can be achieved through the development of advanced intercross lines (AILs), as proposed by Darvasi and Soller (1995). The IBM (Intermated B73 x Mo17) population is an AIL maize population developed by including four generations of random mating following the formation of the F2 generation and prior to the development of inbred lines (LEE et al. 2002). The increased opportunity for recombination has had the effect of expanding the genetic map approximately four-fold compared to non-intermated, conventional, recombinant inbred line (RIL) populations (LEE et al. 2002). The IBM population consists of a relatively large number of lines (302) which have been densely genotyped with more than 2000 molecular markers (COE et al. 2002). Cochliobolus heterostrophus (Drechs.) Drechs. (anamorph=Bipolaris maydis (Nisikado) Shoemaker; synonym=Helminthosporium maydis Nisikado) is a necrotrophic plant pathogen and the causal agent of southern leaf blight (SLB). This disease is frequently found in hot, humid maize-growing areas and was not considered an important pathogen until 1970 when C. heterostrophus race T became prevalent in the US Corn P. Balint-Kurti et al Precise mapping of SLB QTL in maize p5 Belt. Race T was highly pathogenic on Texas male-sterile cytoplasm (cms-T), causing a major disease epidemic in 1970 and 1971 (ULLSTRUP 1972). Since that time, cms-T has been eliminated from elite germplasm and effective polygenic resistance has been introduced. Most of this resistance is quantitative and can be additive or recessive in effect (SCOTT and FUTRELL 1975; LIM and HOOKER 1976; BURNETTE and WHITE 1985; HOLLEY and GOODMAN 1989; BALINT-KURTI et al. 2006) although one qualitative recessive gene, rhm, which primarily conditions resistance in pre-anthesis growth stages, has been mapped to the distal end of the short arm of chromosome six (bin 6.00) (THOMPSON and BERGQUIST 1984; ZAITLIN et al. 1993). The disease, predominantly caused by race O, is still a significant problem in the southern Atlantic coast area of the USA and parts of India, Africa and Western Europe. It has the potential to cause grain yield losses of 40% or more (FISHER et al. 1976; GREGORY et al. 1979; BYRNES et al. 1989). However, use of more resistant germplasm, especially in the USA and western Europe, has largely controlled yield losses due to SLB. To date, three studies have been published on mapping quantitative trait loci for field resistance to SLB in maize (JIANG et al. 1999; CARSON et al. 2004; BALINT-KURTI et al. 2006). In one of these (JIANG et al. 1999) only one SLB QTL was detected and this was observed in only one environment. Another study identified SLB resistance QTL in juvenile maize plants (BALINT-KURTI and CARSON 2006). All of these studies reported QTL spanning large genomic regions. The present study reports SLB resistance QTL identified in the IBM population and compares the results to SLB resistance QTL identified in a conventional RIL mapping population derived from the same parents as the IBM population. QTL identified for time to anthesis are also reported. P. Balint-Kurti et al Precise mapping of SLB QTL in maize p6 MATERIALS AND METHODS Plant Materials: Phenotypic data were collected from the IBM mapping population comprised of 298 F7:8 recombinant inbred lines (RILs) derived from the cross of maize inbred lines B73 (relatively susceptible parent) and Mo17 (relatively resistant parent). This population had been intermated four times at the F2 stage before inbred lines were derived (LEE et al. 2002). Seed of IBM lines was received from the Maize Genetic Stock center and also as gifts from Drs. A. Stapleton and O. Hoekenga. The other population referred to in this paper is a set of 158 F6:7 RILs developed from a B73 x Mo17 cross by selfing directly from the F2 generation by C. Stuber and colleagues (here referred to as the ‘Stuber population’) (CARSON 1998). This population was not assessed in the present study, but previous data derived from it was reanalyzed (see below). Field Trials: All experiments were performed at the North Carolina State University Central Crops Research Station located at Clayton, NC. SLB resistance was evaluated in four environments in this study. Two experiments in separate fields were rated in each of two years, 2005 and 2006. One experiment in each year was artificially inoculated with SLB, and the other was infected solely by natural inoculum. Henceforth these four combinations of treatment and year will be referred to as inoc2005, uninoc2005, inoc2006 and uninoc2006 respectively. Although 298 lines were used in all, due to seed shortage and other factors, all lines were not represented in each environment; 229, 287, P. Balint-Kurti et al Precise mapping of SLB QTL in maize p7 277 and 199 lines were rated respectively in inoc2005, unioc2005, inoc2006 and uninoc2006. Each experiment consisted of two replicates of the IBM population plus parental lines (B73 and Mo17) in complete randomized blocks. With the exception of uninoc2006, experimental units in each case consisted of single-row plots arranged in randomized complete blocks with two replications. Plots were 2 m in length with a 0.6 m alley at the end of each plot. Inter-row spacing was 0.97 m. Twelve seeds per plot were planted in each plot and rows were not thinned, except in the case of uninoc2005, where some thinning was done. Two plots of SLB-susceptible inbred border were planted on all sides of the experiment. Overhead irrigation was applied as needed to ensure satisfactory plant growth. Standard fertilizer and herbicide regimes for central North Carolina were used. For uninoc2006, plot length was 1m and the plants were thinned to about 6 plants per plot. Other details were the same as above. For the Carson 2004 study (CARSON 1998), the Stuber population was inoculated with C. heterostrophus race O, isolate 2-16Bm, in two randomized complete blocks over two years in Clayton, NC. Inoculation techniques used were the same as in the present study. Fungal growth and inoculation: Techniques used for inoculum preparation were identical to those reported previously (CARSON et al. 2004). In the artificially-inoculated experiments, experimental and border plots were inoculated at the fourto six-leaf stage by placing ~20 grains of a sorghum seed culture of C. heterostrophus race O, isolate 216Bm (CARSON 1998) in the leaf whorl of every plant in every plot. After inoculation, P. Balint-Kurti et al Precise mapping of SLB QTL in maize p8 the field was irrigated in order to wet the sorghum seed and allow commencement of fungal growth. Uninoculated experiments were separated from inoculated experiments by