Targeted gene analysis in Ulmus americana and U. pumila tissues

Abstract

Summary Steady-state gene expression was compared between Dutch elm disease (DED)-susceptible Ulmus americana and DED-resistant U. pumila callus, leaf midrib, root and inner bark tissues. Stress-related cDNAs including phenylalanine ammonia-lyase (PAL), chitinase (CHT) and polygalacturonase-inhibiting protein (PGIP) were isolated and compared following RT-PCR of elm tissues. Complete CHT and partial PAL and PGIP cDNA transcripts were identified, each displaying sequence variation between elm species. These transcripts were Dig-labelled and subsequently used for northern analyses of the elm tissues. Midrib and root tissue displayed highest steady-state gene expression compared with inner bark and callus tissues. A modified nucleic acid isolation technique was necessary for downstream RNA analyses. Lithium chloride and polyvinylpyrrolidone were critical for efficient removal of polysaccharides and phenolics associated with some of the elm tissues. Steady-state gene expression is discussed in relation to the tissues investigated. The use of tissues other than in vitro callus culture more closely represents the tissues associated with the elm’s vascular response to DED.