Studies On Specific Proteins And Proteomics Of Flower-bud Induction In Apple Tree

Apple is one of the most important economic fruit trees. There are many factors influencing flower-bud formation and it is necessary for all fruit trees. Recent advances in the conception of flower-bud induction, effects of environmental factors on flower-bud induction, and physiological changes during flower-bud induction are briefly reviewed in this paper. Further investigation is needed to elucidate the molecular mechanism of flower-bud induction. Flower-bud formation specific protein of mature spur from 'Red Fuji', 'Golden Delicious', and 'Red Fuji×Golden Delicious' apple trees (Malus domestica Borkh.) was studied by employing two-dimensional electrophoresis (2-DE), bioinformatics , and mass spectrometry (MS). The results are shown as follows:1. Two-dimensional electrophoresis for apple plant including preparation of protein sample, electrophoresis, and staining was improved to solve the interference with rich polyphenols, tea pigments, and their oxides on the stability and clearness of two-dimensional electrophoresis analysis. About 765 protein spots could be visualized on the 2-DE map by staining. Relative molecular weight of those protein were between 13kD~178kD, most of them were in the range of 13 kD~79 kD. The isoelectric points were found between 4.0~9.5.2. Among varieties 'Red Fuji', 'Golden Delicious', and 'Red Fuji× Golden Delicious', two-dimensional electrophoresis patterns were studied for spurs 3-9 weeks after growth stopped. The result showed that there were 283 differential protein spots existed and changed in quality and quantity on two 2-DE maps for 'Red Fuji' , 'Golden Delicious', and 'Red Fuji× Golden Delicious' at seventh week, of which 162 spots increased and 110 spots decreased in quantity. Four protein spots(16.4, 30.2, 40.3, 65.1kD)were observed the flower bud initial stage1 (inflorescence appears)in two-dimensiononal electrophoresis maps; Three protein spots(39^ 60.2 > 66.3kD)were observed the initial stage 2(Lateral bud appears) in two-dimensiononal electrophoresis maps; one protein spot(77.1kD)was observed the sepal in two-dimensiononal electrophoresis maps.The results can lay down an important foundation in obtaining correlated protein and differential gene in specific period of apple inflorescence development as well as the development mechanism of flower-bud differentiation in molecular level in apple.3. 'RedchieF and 'Red Fuji' apple tree were used as materials to test effect of paclobutrazol (PP333) and GA3 treatment on contents of endogenous hormones in buds. The results showed that PP333 could increase the number of leafy bud nodes, but no effect on number of floral bud nodes. GA3 treatment had no effect on node number of leafy bud on 'Red Fuji', however, it has showed a trend in decreasing the node number of floral bud on 'Redchief' . PP333 and GA3 application made no change on critical period of flower-bud initiation. PP333 enhanced the development of the flower bud and thus shortened period of flower bud formation. GA3 delayed the development of flower bud. The contents of ZR/IAA, ZR/GA3, ABA/ABA and ABA/GA3 ratios were remarkably increased with PP333 treatment. Thus it promoted the flower buds formation. While the contents of ZR/IAA, ZR/GA3, ABA/ABA and ABA/GA3 ratios were decreased by GA3 treatment, it suppressed the flower buds formation.4. Different methods of flowering regulation were used on 'Red Fuji' apple trees. Changes in protein fractions during the flower initiation were determined by IEE-PAGE. The results showed that some of the special proteins were observed in the buds treated with GA3 at the flower induction stage, but disappeared finally. In flower primordium formation stage, intensity of protein bands in flower buds increased with PP333 treatment, and some special proteins were found in flower buds. It is suggested that different methods of flowering regulation might result in different gene expression and characteristics of protein metabolism.5. Four special proteins were observed by peptide mass fingerprinting and matrix-assisted laser desorption ionization time of flight mass spectrometry. By the analysis of four special proteins spots with peptide mass fingerprinting and matrix-assisted laser desorption ionization time of flight mass spectrometry. When Red Fuji apple begin to spur flower buds. At the same time identified proteins and their putative functions were presented through the retrieval different database. One of them, spot No.256(16.4kD)* 298 (30.2 kD) were unknown proteins and spot No.327(40.3kD)was identified as the synthesis enzyme protein. Spot No.367(40.3kD)was identified as a RNA-binding protein involved in transcription. Technical details concerning database queries and successful protein identification rate in the absence of a sequenced genome are reported and discussed.