Study Of Differentially Expressed Proteins During Maize Kernel Development

The development of maize kernel is a process of grain yield and quality formation. Proteins expressed by gene during this process participate in the regulation of different kernel traits. The study of differentially expressed proteins during maize kernel development of 14 inbred lines reveals the molecular mechanisms of kernel development at the protein level and is meaningful for breeding and physiological regulation of grain yield and quality. The differentially expressed proteins during kernel development were studied. The main results are as follows:1. The quality accumulation were cleared by the measured kernel volume, fresh weight, dry matter accumulation, the starch and protein content of the 14 maize inbred lines during kernel development. On the basis of the change curve of kernel fresh weight, volume and dry matter; the kernel development process of the materials were divided into the grain formation period, rapid weight gain period and mature period.2. The quality and two-dimensional electrophoresis result of protein extracted by phenol extraction method. The TCA/acetone method and soluble protein extraction method were compared with Zheng 58 fresh ripe grain as study material. Based on the above results, the soluble protein extraction method was determined as the best for the study of two-dimensional electrophoresis of maize grain protein. The Two-dimensional electrophoresis technology system of maize grain protein was optimized by adjusted parameters of IPG dry strip(24cm, p H4-7), sample volume(900ug), gradient boost voltage, focus 80000 Vhs.3. Express pattern of the identified proteins during the two materials of Zheng 58 and Chang 7-2 were analyzed by Matrix-Assisted-Laser Desorption Ionization/ The time-offlight identified the protein spots differentially expressed 2 times after the atlas of protein two-dimensional electrophoresis of maize grain development were established at 7 days following pollination and at 6 days interval. Expression pattern of 16 proteins including malic enzyme and aldolase etc. are same during grain development of the two maize inbred lines; however, expression patterns of glycine rich proteins were different. About 19 proteins including, triosephosphate isomerase expressed in the whole development process; however, secretory proteins expressed only in a specific period.4. The protein spots differentially expressed in the grain formation period, rapid weight gain period and mature period of kernel development process for all the lines were identified. In total, out of the 107 protein spots; 106 spots were identified effectively and 51 proteins were obtained. A maximum of 20 protein spots were identified as the same protein, and 2 spots for the same protein at least; remaining 30 proteins have specific spots.5. The expression of 51 proteins varies with some expressed in all the lines, while others only in some of the lines. Some of the proteins were expressed in all periods of maize kernel development, while others in one or two periods only. According to the high expression time of the proteins, the proteins were divided into grain formation period, rapid weight gain period, mature period, mid early and mid late period proteins.6. The key proteins affecting the grain yield traits and quality index of maize were determined. The highest correlation with kernel fat content is the hypothetical protein ZEAMMB73-585316 expressed in mature period, and the correlation degree was 0.9501.The second is the putative actin family protein expressed in the grain formation period.The highest correlation with kernel protein content is the hypothetical protein ZEAMMB73-585316 expressed in rapid weight gain period, and the correlation degree was 0.9222, The second is the uncharacterized protein LOC100272498 expressed in the mature period.The highest correlation with kernel starch content is the glycine-rich protein 1 expressed in mature period, and the correlation degree was 0.9323.The second is the globulin 2 expressed in the rapid weight gain period.The highest correlation with kernel dry matter is the alanine aminotransferase expressed in rapid weight gain period, and the correlation degree was 0.9489.The second is the triosephosphate isomerase expressed in the rapid weight gain period.The highest correlation with kernel fresh weight is the class IV heat shock protein expressed in mature period, and the correlation degree was 0.9460.The second is the triosephosphate isomerase expressed in the rapid weight gain period.The highest correlation with kernel kernel volume is the alanine aminotransferase expressed in rapid weight gain period, and the correlation degree was 0.9438. The second is the triosephosphate isomerase expressed in the rapid weight gain period.7. Gene ontology annotation of 51 identified proteins was done using KEGG effectively. 51 proteins are involved in 19 biological pathways, involving 11 cytological components, 8 class of molecular functions. Metabolic pathway analysis showed that 33 proteins were related to 29 metabolic pathways. Biological functions of differentially expressed proteins in the maize kernel development were classified, discussed and analyzed. The functions of proteins such as kernel development, energy metabolism, amino acid metabolism, protein synthesis and metabolism, stress and antioxidant aging, storage protein synthesis and accumulation, hormone and signal transduction, gene transcription regulation, defense related proteins, RNA synthesis and cell activity, gene expression regulation were all successfully identified.8. The mechanism of maize kernel development was preliminarily explored: During the grain formation period, the cell division is strong, and the material metabolism is mainly by the synthesis of carbohydrate and amino acid; During the rapid weight gain period, energy metabolism occupies a dominant position, protein synthesis and amino acid metabolism are also strong, glucose metabolism and purine metabolism is strong and continues to mature perriod, during this period, the body has already started the stress, defense and anti ageing mechanism, at the same time, some proteins unknown, undescribed,or unnamed that are involved in the regulation of developmental processes are expressed in high abundance continues to mature perriod; During the mature period, nutrients accumulated substantially, stress,defense,as well as anti ageing mechanisms are still active, while signal transduction enhanced.