The phosphite responsive transcriptome of phytophthora cinnamomi

摘要: Phosphite has been used to effectively control the soil borne plant pathogen Phytophthora cinnamomi in many horticultural crops, forest trees and natural ecosystems. However, molecular mechanisms behind phosphite action on this are poorly understood. Several studies have shown that inhibits growth zoospore production of P. addition induces significant physiological metabolic changes mycelium. As an approach understanding relevance these pathogen, effect gene expression was investigated using microarray analysis. To construct microarray, RNA extracted from phosphite-treated (40 ug/ml) mycelium isolate MP 80. The chosen concentration inhibited mycelial by 70% but provided sufficient for extractions after 4 days at 25C. The mRNA reverse transcribed into cDNA cloned lambda a library consisting 2 million pfu which 80 % were recombinant phage. inserts sequenced random selection clones library. nucleotide sequences generated revealed range different genes being expressed demonstrated good representation transcripts cinnamomi. types found be included encoding GTP binding proteins involved vesicle transport, structural maintaining cell membrane integrity,elicitors, phosphatases ribosomal proteins. Over nine thousand randomly selected prepared PCR amplification purification construction. Custom made arrays containing 9216 constructed probed with untreated grown medium 40 ug/ml phosphite. Two genes, EF-1 alpha cinnamomin gene, identified qRT-PCR as constitutively also positioned positive controls. In process identifying qRT down-regulated ubiquitin-conjugating enzyme, component ubiquitin/proteasome pathway removal abnormal short lived-regulatory rate limiting enzymes. From further seventy-two altered patterns (fold change > 2) identified. majority spotted array ranging 2- 3.5-fold. Thirty-two up-regulated 16-fold. Characterisation sequencing most highly induced coded ADP-ribosylation factors, ABC cassette transporter glycosyl transferase. A transcript vitamin B6 biosynthesis protein 2.9-fold. contrast, cellulose synthase I, annexin, glutamine synthetase, metallothionein alternative oxidase. results discussed terms possible roles mechanism(s) within P.cinnamomi. This work is first comprehensive screen regulated-gene represents step towards mode organism. This thesis provides valuable information interaction between cinnamomi, future may stimulate discovery novel methods cellular targets pathogenic Oomycetes.