ABSTRACT: Rare sugars are monosaccharides that are rarely present in nature. One of the rare sugars, D-allose had a negative effect on shoot growth of rice. This negative effect to rice growth was not observed in other tested rare sugars. To clarify the response to the D-allose in rice at gene expression level, we performed a microarray analysis using the Agilent Rice Oligo Microarray (44k, custom-made; Agilent Technologies, Redwood City, CA, USA). As a result, treatment of D-allose caused high induction of many defense-related genes including pathogenesis-related (PR) genes in rice. An Agilent Rice Oligo Microarray (44k, custom-made; Agilent Technologies, Redwood City, CA, USA) was used for the microarray analysis. By using RNeasy plant mini kit (Qiagen, Hilden, Germany), total RNA was extracted from shoots of two-leaf stage rice plants that had been treated with 0.5 mM D-allose, 0.5 mM D-glucose, or no sugar for 2 days. For each biological replicate, material from at least five rice plants was pooled to provide a single sample for RNA extraction. We performed 3 biological replicates for each treatment. All microarray procedures and data analyses were performed according to the manufacturer’s instructions. RNA integrity was checked using an Agilent 2100 Bioanalyzer (Agilent Technologies, Redwood City, CA, USA). Total RNA (400 ng) was labeled with Cy-3 or Cy-5 using an Agilent Low RNA Input Fluorescent Linear Amplification Kit (Agilent Technologies). Fluorescently labeled targets were hybridized to Agilent Rice Oligo Microarrays for 17 h at 65°C. After hybridization, following wash processes were performed according to the manufacturer’s instructions, and hybridized microarrays were scanned using an Agilent Microarray Scanner (G2505B and software G2565BA; Agilent Technologies). Feature extraction software (version 9.1; Agilent Technologies) was used to delineate and measure the signal intensity of each spot in the array, and to normalize intensities. The background was measured around each spot as local background, calculated by the Feature Extraction software. Statistical data extraction processes were performed according to the manufacturer’s instructions.
