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Fusarium is the most common flax pathogen causing serious plant diseases and in most cases leading to plant death. To protect itself, the plant activates a number of genes and metabolic pathways, both to counteract the effects of the pathogen, and to eliminate the threat. The identification of the plant genes which respond to infection is the approach, that has been used in this study. Forty-seven flax genes have been identified by means of cDNA subtraction method as those, which respond to pathogen infection. Subtracted genes were classified into several classes and the prevalence of the genes involved in the broad spectrum of antioxidants biosynthesis has been noticed. By means of semi-quantitative RT-PCR and metabolite profiling, the involvement of subtracted genes controlling phenylpropanoid pathway in flax upon infection was positively verified. We identified the key genes of the synthesis of these compounds. At the same time we determined the level of the metabolites produced in the phenylpropanoid pathway (flavonoids, phenolic acids) in early response to Fusarium attack by means of GC-MS technique. To the best of our knowledge this is the first report to describe genes and metabolites of early flax response to pathogens studied in a comprehensive way.

Maize (Zea mays L.) is an important crop that is poorly adapted to cold stress; very little is known about the components of the response to stress. By means of the cDNA subtraction technique, several novel genes, responsive to cold stress, have been identified, including ZmCOI6.1, the function of which is unknown. The predicted ZmCOI6.1 amino acid sequence and its homologue are very similar to proteins in rice and Arabidopsis, suggesting that it belongs to a conserved group of plant proteins. Analysis of the ZmCOI6.1 promoter sequence revealed several conserved stress-responsive cis-acting elements. Further characterization of expression showed that ZmCOI6.1 was induced not only by cold, but also by drought and salinity as well as by the signaling molecules abscisic acid and salicylic acid, indicating that ZmCOI6.1 is a stress-responsive gene. ZmCOI6.1 is alternatively spliced and yields two transcripts, the level of which change depending on the stress, indicating a possible mechanism of regulation at the splicing level. Constitutive expression of ZmCOI6.1 in Arabidopsis yielded plants that were less tolerant to abiotic stress, providing evidence that ZmCOI6.1 may be a negative regulator.

The plant regulator methyl-jasmonate (MeJA) mediates diverse developmental processes and defense responses which induce a variety of secondary metabolites. This study reports the regulatory network of MeJA-induced genes in adventitious root cultures of the medicinal herb Bupleurum kaoi. A cDNA microarray containing 465 unique expressed sequence tags (ESTs) as probe sets, generated with a PCR-Select cDNA Subtraction Kit, was employed to profile the transcriptome of MeJA induction at thirteen points in time. Real-time PCR was performed to verify changes in expression of 36 unique ESTs (uniESTs), detected via microarray. Genes upregulated by MeJA include those involved in saikosaponin, amino acid, phenylpropanoid and jasmonate biosynthesis, defense signaling, plant hormone regulation and stress protection; indicating that the JA signaling pathway interacts with other signaling pathways leading to transcriptional reprogramming in B. kaoi adventitious roots. Some of these MeJA-induced genes have applications in agricultural, pharmaceutical and food industries.

Two subtracted cDNA libraries were constructed by reciprocal subtractive hybridisation between sugarcane immature (low sucrose-accumulating) and maturing (high sucrose-accumulating) internodal tissue. The subtracted libraries contained high, moderate and low abundance transcripts. To isolate cDNAs differentially expressed during culm maturation, 400 random clones (200 from each library) were systematically arrayed onto nylon filters and screened with total cDNA probes prepared from immature and maturing culm poly (A) + RNA. Results indicated that 36 and 30% of the total number of cDNAs analysed were preferentially expressed in the immature and maturing culm, respectively. Northern analysis of selected clones confirmed culm developmental stage-specific and -preferential expression for most of the clones tested. Expressed Sequence Tags (ESTs) generated by partial sequence analysis for all 132 differentially expressed clones indicated 95 unique transcripts. Partial sequence information could assign putative identities to 66% of the differentially expressed ESTs. The majority of ESTs with a putative identity were homologous to genes associated with cell wall metabolism, carbohydrate metabolism, stress responses and regulation, where the specific ESTs identified in the immature and maturing culm were distinct from each other. No developmentally regulated ESTs directly associated with sucrose metabolism were detected. This suggests that growth and maturation of the sugarcane culm is associated with the expression of genes for a variety of processes. This study demonstrates that a combination of cDNA subtraction with macroarray screening is an effective strategy to identify and analyse candidate developmentally regulated genes in sugarcane.