| Pear?Pyrus?usually blooms a huge amount of flowers when it grows in nature,so it needs auxiliary artificial thinning for assuring harvested fruit's quality,comparing with labor,chemical thinning saved time and thinned timely.However,the commonly used chemical thinning agents in orchard production management have more or less side effects,such as environmental pollution or reducing fruit quality,5-ALA as a new chemical thinning agent which is environmental friendly and can improve the fruit quality arises at the historic moment.Our previous study found that 5-ALA had the potential of thinning,but the effective concentration and its role was not clear.In order to determine the thinning effect and effective concentration of 5-ALA,this paper has done a series of experiments in field and in vitro to explore,meanwhile,we studied the thinning mechanism of 5-ALA from a microscopic level with optical microscope,fluorescence microscope and transmission electron microscope?TEM?,the main results are as follows:1.Spraying 150 mg·L-1 5-ALA in full bloom could significantly reduce the fruit setting rate of 'Mingshui' pear,and then met the needs of production,meanwhile,5-ALA could significantly improved soluble solid content and soluble sugar content of mature pear fruit,so the fruit quality was improved.The results of pollen germination in situ test showed that sraying 20 mg·L-1 5-ALA at 2,8,12,24 h before artificial polliation could effective inhibit the growth of pollen tube in the style,so the inhibition of pollen tube elongation was an important biological basis of 5-ALA's thinning effect.2.20,30,40 mg·L-15-ALA solution could significantly reduce the germination rate of pollen and the length of pollen tube.Meanwhile,observing by optical microscopy,we found that 5-ALA in the culture medium caused abnormal growth of pollen,for example,the color of pollen tube tip deepened,the top end of the transparent zone was narrowed or disappeared,pollen tube tip or near tip became swelled and deformed,burst,even internal flowed,etc.These abnormal phenomena showed that 5-ALA had a significant effect on the normal growth of pear pollen.3.Adding 40 mg·L-1 5-ALA in the culture medium significantly increased the number of vesicles in pollen tubes;Secondly,adding 20,30,40 mg·L-1 5-ALA in the culture medium,the fluorescence brightness of callose in pear pollen tube cell wall structures increased obviously.At last,we studied the interaction between 5-ALA and microtubule stabilizer taxol during pollen germination and pollen tube elongation via adding them in control based culture medium.The study showed that low concentration presented the pear pollen tube elongation length growth,and high concentration shorted it.Among them,when the concentration of paclitaxel was 0.1?g·mL-1,the effect on promoting pear pollen tube elongation growth effect was the most significant.Then,adding 20,30,40 mg·L-1 5-ALA also was added after it,we analyzed statistically the pollen germination rate and pollen tube length,taxol could relieved the negative effect of different concentrations 5-ALA on pear pollen tube length.At the same time,0.1 ?g·mL-1 taxol could significantly relieved the inhibition effect of 5-ALA on pear pollen tube length,it was showed that 5-ALA could destroy the structure of pollen tubes.In summary,5-ALA could destroy the stability of the tube in the tube,broken the balance of the vesicle transport and the normal construction of the cell wall.On the one hand,the number of vesicles in the tip of pollen tube was significantly increased,and the vesicles became smaller,The rapid increasing of vesicles formed large vesicle clusters in limited space of the pollen tube.On the one hand,the vesicles which were near the pollen tube wall had been fused into one piece,the abnormal vesicle clusters extruded the cell wall of pollen tube to make it expanded and deformated;On the other hand,the vesicles in the tube clustered together to surround the organelles,for example,the mitochondria,so that its structural deformated and function was blocked.Besides,the results of the fusion of vesicles and vacuoles in the tube led to the rupture of the vacuole membrane and the vesicle membrane,then harmful substances in the bubble were out,pollen tube tip cytoplasmic density became larger,this willed not only damage the normal osmotic potential in the tube but also hinder the normal physiological and biochemical reactions.On the other hand,callose in wall increased,the wall inside appeared a phenomenon of callose deposition,then pollen tube cell wall's rigidity enhanced,ductility weakened,it not only slowed down the growth rate of pollen tube elongation,but also affected the normal endocytosis of pollen tubes.Above phenomenon caused the abnormal growth of pollen,resulting in a significant reduction in pollen germination rate and pollen tube elongation growth length,and prevent the occurrence of fertilization,resulting in thinning effect. |
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