Comparative Analysis Of The Protein Expression At Different Developmental Stages Of Microspore In Shaannong138 Wheat

Wheat is the most important crop in the world, which is also the major food crop in China. The research and utilization of wheat anther culture was an important way to improve the efficiency of wheat breeding, However, the induction rate of callus and seeding rate were very low in the process of anther culture, which was the bottleneck in the development of anther culture. the basic study on the anther culture ability of wheat should be strengthen, In order to service agricultural development by making better use of wheat anther culture technology.The anther culture ability of wheat is the anther callus in the process of anther culture and the differentiation rate of green plantlet, which is affected easily by many factors, such as the genotypes of material, the developmental stage of anther microspore, media, culture conditions and so on. The developmental stage of anther microspore is very important to the induction rate of callus of anther culture in these factors. Many results showed that the mid and late uninucleate pollen was the best vaccination period in anther culture in vitro of wheat, in which the induction rate of callus of anther culture was the highest. At present, the induction rate of callus was calculated by comparative test of anther culture in the previous studies, which the reason could not be explained, so the reason that the induction rate of callus of anther culture was the highest in the mid and late uninucleate pollen and the mechanism of protein metabolism in the developmental stage of anther were explored in this paper, in order to find the protein that had the relation with the anther culture ability of wheat in this stage.In this paper, the whole proteome at the early uninucleate pollen period, mid and late unimucleate pollen period and binucleate pollen period of microspore development was systematic studied by proteomics technology with the Shaannong138 as the material.The main results and discusses were as follows:1. The total protein bands at the early uninucleate pollen period, mid and late unimucleate pollen period and binucleate pollen period of microspore development were analyzed by SDS-PAGE electrophoresis in this paper. The result showed that two protein bands appeared at the binucleate pollen period of the microspore development, whose molecular weight of 50.7kDa,48.2kDa, and these two bands appeared alone at the early and mid-late uninucleate pollen period respectively.2. The total protein bands at the mid and late unimucleate pollen period and binucleate pollen period of microspore development were analyzed by two-dimensional electrophoresis technology in this paper. The result showed that:(1) There were about 450 clear protein spots could be detected between the isoelectric point (pI)4 and (pI)7, in which there were 26 different protein spots. There were 11 different protein spots in the mid and late unimucleate pollen period, in which there were 5 spots disappeared and 6 spots significantly reduced from the mid and late unimucleate pollen period to binucleate pollen period. And there were 15 different protein spots in the binucleate pollen period, in which there were 5 spots re-expressed and 10 spots significantly enhanced in this stage. (2)The proteins that expressed in two-dimensional and one-way electrophoresis during the two stages were very similar according to the distribution of MW, and the difference of protein spots was extremely significant, which proved the reliability of one-way electrophoresis results.3. The peptide fingerprinting (MALDI-TOF-MS) of 14 different protein spots were analyzed by matrix-assisted laser desorption time of flight mass spectrometry and using Mascot to search in the NCBInr database, which identified 11 protein spots, however, there were 3 protein spots could not be identified.1 protein spot was identified as UDP-glucose-1-phosphate uridylyltransferase, chlorophyll a-b binding protein, pentatricopeptide repeat protein PPR566-6, cysteine proteinase inhibitor A and ubiquitin; 2 protein spots were identified as S-adenosyl-L-homocysteine hydrolase, two hypothetical protein and two stocking-increasing protein. The functions of proteins identified were related to metabolic process of material energy, carbohydrate metabolism, protein degradation and cell defense, which were likely associated with the anther development of wheat.