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Cloning and characterization of a rhamnogalacturonan hydrolase gene from Botrytis cinerea
H J Chen1, D L Smith, D A Starrett
1Horticultural Crops Quality Laboratory, USDA, Agricultural Research Service, Beltsville, MD 20705, USA.
Abstract:
Rhamnogalacturonan hydrolase (Rgase A) cleaves alpha 1--2 linkages between rhamnosyl and galacturonosyl residues in pectin. A 1.9 kb RGase A cDNA clone (BCRHGA) was isolated from a B. cinerea cDNA library using a PCR-amplified Aspergillus aculeatus RGase A probe. It's 1.7 kb open reading frame had 62% identity at the amino acid level with A. aculeatus RGase A. Northern blots of B. cinerea total RNA probed with BCRHGA revealed a 2 kb band, suggesting the cDNA clone is full or nearly-full length. To determine mRNA expression of the gene, B. cinerea was grown in media containing 0.5% apple pectin, 0.5% rhamnogalacturonan-I and 1% glucose carbon sources. Northern analysis revealed the BCRHGA gene was expressed on all carbon sources, but with different patterns of expression. B. cinerea RGase A appeared to be coded for by a single or low copy number gene based on Southern analysis.
