Proteomic Identification,Cloning And Verification Of Key Genes Associated With Nitrogen Utilization Efficiency In Cotton

To identify key genes, Jinmian 11 and Simian 3 were used as materials, iTRAQ quantitative proteomics technology was applied to study the protein expression of cotton root under low nitrogen stress in this paper. The key genes associated to efficient utilization of nitrogen were identified based on the systematic, comprehensive and accurate comparison combined with bioinformatic analysis. Real time RT-PCR technology was applied to verify the expression of these candidate genes. In current study were explored the molecular mechanism of cotton efficient utilization of nitrogen and two genes were cloned for cotton breeding of efficient utilization of nitrogen. In this paper, the main results are as follows:1. The iTRAQ quantitative proteomics technology was employed to screened differentially expressed proteins in root of two cotton cultivars under low nitrogen stress. Total 1045 differentially expressed proteins were identified, and 178 up-regulated proteins in both two cotton varieties. Total 46 genes were initially chosed as key genes associated with efficient utilization of nitrogen in cotton.2. GO analysis showed that about 47% of the identified proteins are related to the metabolic processes and about 38% of the protein are associate with the cellular processes according to their assortment on biological process. According to their classification of molecular function, nearly 44% of the identified protein have catalytic activity,40% of the protein have binding activity, suggesting that almost all identified proteins have functional activity.3. The pathway enrichment analysis of the identified protein showed that 4 identified protein including ferredoxin-nitrite reductase, carbonic anhydrase, glutamate dehydrogenase (NAD (P)+) and asparagine synthase are up-regulated in Nitrogen metabolism pathways.4.35 differentially expressed proteins genes were checked by qRT-PCR. The results showed that the expression of 19 genes including prep, pdf, popt, syds, csh, gly, hpp, mettp, etlfa, pdp, upl, arp, cgs, glu, nrp, alip, pgp, prp4 and TO at mRNA level are consistent with those that at protein level. While the other 16 gene showed inconsistent expression results.5. Two genes including cgs and TO that closely are related to nitrogen efficiency were cloned in the paper.