Identification Of Differentially Expressed Genes By CDNA-AFLP Approach During Water Stress In Wheat Leaves

Drough is an environmental stress facror which has the most impact on crop productivity, yield losses caused by it is more than the sum of the loss by other stress factors. The drought–resistant trait of plant is controlled by many genes, and influenced by genetic and other factors. To some extent, drought tolerance of crops have similar molecular basis of the expression of many genes induced by water stress and water stress resistance in crops. How to improve the drought resistance and productivity of crops under drought stress has been the research focus. It is worthful to find the molecular mechanism of drought-resistant crops for crop breeding and cultivation techniques .In our research Xinchun6 was used as the experimental material, it is a typical drought- resistant and a high water-use-efficiency variety. The gene expression differences betwenn water deficit treatment and control were analyzed by using cDNA-AFLP technique. A corresponding research system was established based on gene expression differences in wheat leaves and we found gene expressed sequence tags which may be relative to the efficient use of water and drought resistance.The results are as follows:(1)We found the best method to obtain total RNA of wheat leaves for cDNA-AFLP by comparision.(2)By comparing two sets of cDNA-AFLP program, we determined that the double digeste cDNA-AFLP method is reliability and better than the single digest cDNA-AFLP method. So the double digeste cDNA-AFLP method is more suitable for our research.(3)Gene expression of cDNA-AFLP technology system for wheat were established and the PCR system was optimized.(4)The gene expression differences of Xinchun6 by water deficit were analyzed using 253 pairs of selective primers, we found 748 different bands, and 44 of them were recovered, these differences were manifested as bands up different types.(5)The 10 differential bands were selected to clone and sequence, and the differentially expressed fragments were analyzed using BLAST. 10 sequences were all found in the BLAST database, mainly related to the categories including signal transduction, transcription regulation, energy metabolism, cell growth and development, stress responses. 1)The growth and development of water stress response genes, which mitochondrial 26rRNA have homology higher TDFHh2-1,TDFHh4-3 and TDFHh3-3; And chloroplast DNA sequence of high homology of TDFHh7-1. 2)The response to water stress in cell metabolism genes: NADH dehydrogenase and the main 7 subunit and pyruvate kinase homology higher TDFHh15-2 and TDFHh16-2. 3)The response to water stress response and defense genes: mainly have homology higher cytochrome P450 and retrotransposons homologous mRNA the TDFHh1-3 and TDFHh6-4. 4)The function-unknown of water stress response genes: the mRNA sequence of Zea and hypothetical protein homology higher TDFHh17-3 and TDFHh21-1.