2006. Finemapping, cloning, verification, and fitness evaluation of a QTL, Rcg1, which confers resistance to Colletotrichum graminicola in maize

2006. Finemapping, cloning, verification, and fitness evaluation of a QTL, Rcg1, which confers resistance to Colletotrichum graminicola in maize

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ABSTRACT

Colletotrichum graminicola (Cg) causes anthracnose stalk rot and anthracnose leaf blight of maize and other graminaceous hosts (Bergstrom and Nicholson, 1999). Anthracnose has been found to occur in the United States since 1855 and occurs in the Americas, Europe, Africa, Asia, and Australia. Over 37.5 million acres are infested annually in the United States with average yield losses of 6.6%. Previously, a QTL region on chromosome 4 was reported to carry resistance to Cg in the maize line DE811ASR (Jung et al. 1994). DE811ASR was further backcrossed to DE811 (susceptible maize line) to develop DE811 Rcgl which was used to develop a BC7 segregating population for mapping. Utilizing 4784 genotyped BC7 individuals, 856 BC7 individuals were selected as recombinants within the region of interest. The selfed progeny of those 856 individuals were phenotyped to obtain family means, and a high-resolution genetic-linkage map was constructed to further resolve the previously described QTL. Initial markers identified two physical contigs within the region of interest. Using public and private physical and genetic integrated maps, the gap between the contigs was closed, and a complete tiling path through the region was constructed. Additional PCR-based fragment length polymorphic markers were created from BAC end sequences, overgoes and ESTs present on the BAC clones. Subsequent analysis of phenotypic data integrated with the BC7 genetic-linkage map resolved the QTL location to an approximate 3 cM region (~400kb physical distance based on the B73 and Mo 17 physical maps) with a peak LOD score of approximately 55. This 400kb region was sequenced using the corresponding BACs from a library made from the resistant parent, and a candidate gene for resistance to Cg was discovered in the region. Four independent Mu insertion events were identified in putative susceptible mutants derived from a targeted Mu-transposon population containing the resistance gene. Sequence data confirmed the presence of the Mu element within the candidate gene causing these plants to be susceptible to Cg.

The effects of Cg on maize growth and development can be very substantial; estimates of yield loss range from zero to over 40%. A multi-year multilocation trial was conducted to evaluate the agronomic and disease performance of DE81 IRcgl isohybrids compared to DE811 isohybrids when inoculated with Cg. Isogenic hybrids created by crossing each isogenic line, DE811 and DE81 IRcgl, to a diverse set of inbred lines, H99, Mo 17, B73, LH132, and DE4 were evaluated in a randomized complete block design over three locations in 2004 and four locations in 2005 across Delaware with three replications per location. A more detailed split-split plot design was conducted in 2005 to determine if any fitness costs were associated with the isogenic line DE81 \Rcgl when inoculated with Cg, wounded (inoculated with water), or not inoculated. The isogenic hybrids were created by crossing DE811 and DE81 \Rcgl with Mo 17 and B73 and planted in six locations across the U.S. with three replicates at each location. The data indicate that the near isogenic line DE81 \Rcgl does not cause significant negative effects in hybrid combinations for most agronomic traits compared to hybrids made with the recurrent parent, DE811. The results suggest that there was no apparent fitness cost associated with Rcgl in uninoculated or wounded treatments. The presence of Rcgl reduced the overall economic impact incurred by infection in inoculated plots by almost three-fold in comparison to isohybrids that lacked the gene. The resistance appears to act on delaying the onset of the disease until later in the season when the pathogens effects have less impact on plant yield. These results in addition to the asexual life cycle and limited variability of Cg, suggest that Rcgl may be a very durable disease resistance. These data also indicate the importance of diversity and germplasm conservation and evaluation. The low frequency of Rcgl in modem maize hybrids indicates that this resistance may have been lost without a sustained effort to maintain and evaluate diverse germplasm .