Microarray analysis of endophyte-infected and endophyte-free tall fescue

Abstract

Many grasses have mutualistic symbioses with fungi of the family Clavicipitaceae. Tall fescue can harbor the obligate endophyte, Neotyphodium coenophialum that is asexually propagated and transmitted via host seeds. Total RNA was isolated from pseudostems of known endophyte-infected (E+) and endophyte-free (E−) plants and tested in triplicate on the Affymetrix Wheat Genome Array GeneChip ® and Barley1 Genome Array GeneChip ®. Overall 14–15% and 17–18% of the probe sets were called present on the wheat and barley chips, respectively. In order to identify genes that were specifically differentially expressed between the E+ and E− tall fescue, a combination of both barley and wheat target sequences that were differentially expressed (greater than twofold) that were similar on both chips on both barley and wheat arrays yielded 32 probe set (genes) that were differentially expressed. Tall fescue ESTs were identified for a number of the probe sets that were differentially expressed on the barley and wheat arrays. PCR primers were designed to fescue ESTs and tested to verify the expression profile observed in the microarray experiments. Some primers confirmed the expected results, although in other cases no differences were observed between the E+ and E− plants, or the results were contrary to what was expected. Our results suggest that while some differentially expressed genes were identified by this method, the cross-species hybridization appears to have significant limitations for the transcriptome analysis of tall fescue.