Characterization and expression of high temperature stress responsive genes in bread wheat (Triticum aestivum L.)

To elucidate the effects of high temperatures, wheat plants (Triticum aestivum cv. CPAN 1676) were given heat shocks at 37°C and 42°C for two hours, and responsive genes were identified through PCR-Select Subtraction technology. Four subtractive cDNA libraries, including three forward and one reverse subtraction, were constructed from three different developmental stages. A total of 5500 ESTs were generated and 3516 high quality ESTs were submitted to Genbank. More than one third of the ESTs generated fall in unknown/no hit categories upon a homology search through BLAST analysis. A large number of high temperature responsive genes have been identified and characterized. Reverse subtraction analysis in developing grains showed extensive transcriptional changes upon heat stress as revealed by comparative analysis with forward subtraction. Differential expression was confirmed by cDNA macroarray and by northern/RT-PCR analysis. Expression analysis of wheat plants subjected to high temperature stress, after one and four days of recovery, showed fast recovery in seedling tissues. However, recovery was small in the developing seed tissue after two hours of heat stress. Ten selected genes were analysed in further detail by quantitative real-time PCR in an array of 35 different wheat tissues representing major developmental stages as well as different abiotic stresses. Tissue specificity was examined along with cross talk with other abiotic stresses and putative signalling molecules. The results obtained contribute towards understanding the regulation of genes at different developmental stages in wheat crucial to withstanding and recovery from heat stress.