Nitrogen Efficiency Of Genotype Selection And Related Gene Cloning In Cotton

Object:Cotton is one of the most important economic crops in Xinjiang. Nitrogen plays a important role in the cotton growth processand lots of fertilization and low efficient utilization in improving cotton output, but also bring the production costs, soil resources degradation and environmental pollution, a series of agricultural production and the ecological environment problems. Therefore, screening nitrogen efficient utilization of cotton varieties (strains), and using different genotypes uptake of biological characteristics analysis the physiological and biochemical mechanism of cotton and fully exploit nitrogen efficiency of genes. Cultivation of efficient utilization of nitrogen to improve cotton varieties of nitrogen efficient utilization, conservation of resources, reduce and control environmental pollution, finally realizes the sustainable development of agriculture has the important practical significance.Methods:The experiment with nitrogen efficiency of cotton varieties (Xinluzao 12) and use the cDNA-AFLP technique to combine with bioinformatic approaches, clone cotton nitrogen efficient utilization related gene, use bioinformatic method to sequence alignment and functional preliminary forecast for further research foundation of the gene function.Results:1. This experiment is conducted to study the growing and Nitrogen (N) nutrition and to screen of the nitrogen high efficiency of 33 cotton varieties through water culturing. The experiment adopted H-CAB ingredient water culturing and installed two N levels (1 mmol·L-1 and 5 mmol·L-1),and every level is three repetitions, analytic the total plant dry weight, N utilization index of total plant, N concentration of plant, the total plant weight, root-top ratio of different cotton varieties. Significant positive correlations (r=0.946**, N=33, P<0.01) were observed between the relative dry weight of the whole plant with N concentration of plant, and positive correlations (r= 0.432*, N=33, P<0.05) were observed between with N utilization index of total plant. Thus, the total plant dry weight could be first screening index for low N tolerance. Positive correlations (r= 0.445*, N=33, P<0.05) were observed between N concentration of plant with N utilization index of total plant, and they could be secondary screening index for low N tolerance. The results indicated that there were great differences among cotton varieties in the low nitrogen ability, XLZ12, XLZ21, XLZ26, XLZ31, XLZ32, XH25 had higher N utilization efficiency, stronger N absorption ability, and higher N utilization index in low N level; while cultivars PTYH, XLZ 19, XH3, XH13, AJJL and XLZ 36 had lower N efficiency, fewer N absorption ability and lower N utilization index in normal N level.2. In this study, cDNA-AFLP technique was used to screen out N efficient genotypes Xinluzao 12 related genes from N stress tolerance of cotton material by using 64 pairs of primers. 122 differently expressed TDFs were obtained,51 of them were cloned, sequenced and the sequences were analyzed. The result showed that 22 TDFs have homologous genes on NCBI,7 TDFs have photosynthesis function,6 TDFs have possible signal transduction function,4 TDFs have membrane permeability function,1 TDFs have possible Cell metabolism function,4 TDFs have Ion conversion Function. They have highly homology with populus trichocarpa and phytochrome and disulfide isomerase and transglycosylase.3. Bioinformatic and RT-PCR are used to clone disulfide isomerase of full-length related with N utilization, this cDNA colne was 1484bp,5'coding region is 82bp,3'coding region is 322bp, and its largest open reading frame encoded 360 amino acids. The gene was designated as GhPDI (GenBank accession:ES802670.1). Take the gene of homology comparison in NCBI, find have highly homology with ricinus communis is 88%. For protein structure prediction show that the PDI genes encoded protein typically the PDI gene families of protein degradation pathway (ERAD), protein transport pathway, possible judge the preliminary PDI genes for cotton.Conclusion:This study used plant nutrition and molecular biology, combined with cotton nutrition characters from improved,33 cotton varieties (or lines) of nitrogen efficiency genotype selection at seedling, on this basis, using cDNA-AFLP difference display technology combined with bioinformatic approaches from cotton nitrogen efficiency of varieties (xinluzao 12) in a cloning under low the two Two nitrogen key priority express the length of heterogeneous enzyme gene sequences, and using cDNA bioinformatics method sequence alignment and functional preliminary forecast.1. The results indicated that there were significant differences among cotton varieties in the N utilization index through screening, analysis relative dry weight of the whole plant with N concentration of plant and N utilization index, could screening cotton nitrogen efficiency simple and fast form large cotton varieties germplasm resistance of preliminary.2. Many genes about cotton have changed, such as signal transduction and photosynthesis, using cDNA-AFLP difference display technology detach the gene in low N concentration can lay the foundation through study the low nitrogen molecules cotton mechanism and use genetically modified cotton varieties.