Direct screening of a small genome: estimation of the magnitude of plant gene expression changes during adaptation to high salt

Abstract

Mesembryanthemum crystallinum (common ice plant), a facultative halophyte with a genome size of 393,000 kb, was used to estimate the magnitude of changes in gene expression in response to environmental stress by excess salt. Such treatment induces a water-conserving pathway of carbon assimilation (CAM) which is, at least in part, transcriptionally controlled. From a genomic library, 200 phage containing approximately 3200 kb (0.8% of the genome) were randomly selected. The inserts in these clones could be divided into four classes ranging from highly repetitive DNA (class I clones) to single-copy DNA (class IV clones). The inserts of the 166 clones of classes II to IV were digested with various restriction enzymes and the fragments were analyzed by hybridization with radioactively labelled mRNA isolated from stressed and unstressed leaves. We found that a total of approximately 140 DNA fragments hybridized with the RNA probe. Among those, several differentially regulated transcripts were observed. Stress-dependent fluctuation of mRNA abundance was verified by Northern analyses: one mRNA, not detectable in unstressed leaves, appeared in stressed leaves, while steady-state levels of three transcripts decreased during stress. All regulated signals are derived from low abundance mRNAs, which may be missed during screening of cDNA libraries. We conclude from these results that, for the entire genome, on the order of more than one hundred genes are differentially regulated in response to salt stress.