Xun Wang

Gene Expression Phenotypes of Arabidopsis Associated with Sensitivity to Low Temperatures

Chilling is a common abiotic stress that leads to economic losses in agriculture. By comparing the transcriptome of Arabidopsis under normal (22°C) and chilling (13°C) conditions, we have surveyed the molecular responses of a chilling-resistant plant to acclimate to a moderate reduction in temperature. The mRNA accumulation of approximately 20% of the approximately 8,000 genes analyzed was affected by chilling. In particular, a highly significant number of genes involved in protein biosynthesis displayed an increase in transcript abundance. We have analyzed the molecular phenotypes of 12 chilling-sensitive mutants exposed to 13°C before any visible phenotype could be detected. The number and pattern of expression of chilling-responsive genes in the mutants were consistent with their final degree of chilling injury. The mRNA accumulation profiles for the chilling-lethal mutants chs1, chs2, and chs3 were highly similar and included extensive chilling-induced and mutant-specific alterations in gene expression. The expression pattern of the mutants upon chilling suggests that the normal function of the mutated loci prevents a damaging widespread effect of chilling on transcriptional regulation. In addition, we have identified 634 chilling-responsive genes with aberrant expression in all of the chilling-lethal mutants. This reference gene list, including genes related to lipid metabolism, chloroplast function, carbohydrate metabolism and free radical detoxification, represents a potential source for genes with a critical role in plant acclimation to suboptimal temperatures. The comparison of transcriptome profiles after transfer of Arabidopsis plants from 22°C to 13°C versus transfer to 4°C suggests that quantitative and temporal differences exist between these molecular responses.

Gene Expression Microarrays: Improvements and Applications towards Agricultural Gene Discovery

In agriculture, the associations between genes and resulting traits revealed by high throughput approaches such as transcription profiling could be used to select more environmentally friendly chemicals for plant protection and to develop plants with increased grain yields and better nutrition value, with more resistance to diseases and tolerance to abiotic stress. However, one of the major challenges to apply such approaches is the limited genomic information for most of the very diversified crop species. We developed multiple strategies and platform technologies to address this issue. Here we report our improvements of these technologies towards large-scale transcription profiling and their applications in agricultural gene discovery.

Differential gene expression analysis by micro-preparative capillary gel electrophoresis

Differential display analysis by cDNA fractionation, collection of differentially expressed fractions of interests and their downstream characterization is demonstrated. cDNA pools from two strains of Cochliobolus heterostrophus fungus were generated by specific restriction digestion and selective ligation. Micropreparative separation and isolation of differentially expressed transcript representatives were accomplished by high-performance capillary gel electrophoresis. The collected individual DNA molecules were polymerase chain reaction amplified and sequenced to create expressed sequence tags for the genes of interests. High resolving power and sensitivity of capillary gel electrophoresis enabled fast and automated processing of minute amounts of cDNA samples with high precision.