Isolated Identification And Function Of Specific Proteins In Grafting Healing Process Of Carya Illinoinensis

Pecan(Carya illinoinensis) is one of the famous economic forest tree species in the word. In this experiment, the patch budding grafted unions of pecan at its different development stages were used as materials. We explored the optimal protein extraction method in pecan. The suitable and efficient two-dimensional electrophoresis system, the protein expression profiles of grafted unions at its different developmental stages were established and constructed. Differential proteins related grafting survival were filtrated, identified by mass spectrometry and Studied. We aims to provide theoretical basis and technical support for revealing healing mechanism of pecan grafted unions. The main results are as follows:1. An optimal protein extraction method and efficient two-dimensional electrophoresis system was established. The Tris-HCL extraction method was optimal, in which two-dimensional electrophoresis was conducted with immobilized pH4-7 IPG strips and 260?g sample.2. The 2-DE protein maps of different development stages in the grafting healing process of pecan were analyzed, the general numbers of protein increased then decreased then increased during development of the grafted unions. The period that had quite different protein numbers were between 0 to 3 days after grafting, between10 to 15 days after grafting, between 25 to 30 days after grafting. 138 differentially expressed proteins were detected. These proteins mainly concentrated in day 3, 10, 15, 25, 30 after grafting. It indicated the physiological healing of pecan grafting was active during this period. At the third day after grafting, the grafted union could began to generate callus. At 10 to 15 days, the vascular cambium was formed. At 25 to 30 days after grafting, vascular bundles was formed.3. The 121 spots of differential protein during six critical healing periods(1, 3, 6, 10, 15, and 25 days after grafting) were selected and analyzed by mass spectrometry(MS), 110 of them were identified. The 110 spots concentrated in pH 5-7, 20-60 kDa molecular weight scope. They were classified to nine functional classes, in which 43 proteins were involved in energy metabolism(39.09%), 8 proteins were involved in secondary metabolism(7.27%), 5 proteins were involved in protein synthesis and metabolism(4.55%), 8 proteins were involved in amino acid metabolism(7.27%), 1 protein was involved in cell metabolism and cycle regulation(0.91%), 3 proteins were involved in signal transduction(2.73%), 7 proteins were involved in transcription factor(6.36%), 5 proteins were involved in functional regulation(4.55%), 16 proteins were involved in stress response and defense(14.55%) and 14 functional proteins(12.73%) were unknown.