Analysis On The Relationship Between Physiological Fruit Drop And Content Of Four Endogenous Hormones In Different Parts Of Fruit In Citrus

Citrus is one of the most important fruit trees in China,which belongs to Rutaceae,Aurantinoideae.In 2017,citrus yield reached about 38.168 million tons,second only to apple.Citrus was easy to flower,but a lot of flower and fruit will drop at early stage,especially for seedless or seed-few varieties,their early fruit drop and pre-harvest fruit drop were more serious.Parthenocarpy ability was an important factor affecting the yield of seedless citrus,which has a great influence on cultivation measures and management costs.Endogenous hormones plays an important role in processes as plant flowering and fruiting,maturity and senescence.To explore the relationship between endogenous hormones in different fruit parts and physiological fruit drop and parthenocarpy will provide a light on both citrus breeding for varieties with strong parthenocarpy ability,fewer physiological fruit drop,and the use of exogenous plant growth regulator to enhance the flowering and fruit setting or thinning.In this study,endogenous hormones were measured by HPLC in different parts of pollinated and non-pollinated fruits of W-murcott tangor which has a strong facultative parthenocarpic ability,and of citrus varieties showing different degree of pre-harvest fruit drop.Meanwhile,the ovule and fruit development as well as physiological fruit drop process were observed.The main research results are as follows:1.Optimized and established of endogenous hormones extraction and detection methodsThe extraction method of liquid-liquid extraction and HPLC detection condition of zeatin(ZT),gibberellic acid(GA3),auxin(IAA)and abscisic acid(ABA)were optimized.The standard recovery rate of ZT,GA3,IAA and ABA were above 80.9%,and the linear correlation coefficients of endogenous hormones regression equation were above 0.999 with the extraction method.The target substances in the sample were separated well with a sharp peak and without towing peak under the optimized detection conditions.The extraction method and detection condition were optimized as follows:(1)Extract liquid:80%methanol(including 1%acetic acid) Extraction tool:50 mL centrifuge tubes Extraction order:The acid endogenous hormones GA3,IAA and ABA were extracted first,and then the weakly alkaline endogenous hormone ZT was extracted by adjusting PH.(2)Mobile phase:0～3 min,V(methanol):V(acetonitrile):V(phosphate buffer)=5:5:90;3～10 min,V(methanol):V(acetonitrile):V(phosphate buffer)=15:15:70;10-15 min,V(methanol):V(acetonitrile):V(phosphatebuffer)=30:30:40;15-20 min,V(methanol):V(acetonitrile):V(phosphate buffer)=20:20:60.Injection volume:10 μL Column temperature:30 C Detection wavelength:ZT、GA3、IAA were 210 nm,ABA was 254 nm.2.Analysis on the relationship between T value and physiological fruit dropThrough the detection and analysis of endogenous hormones in different citrus varieties during physiological fruit drop,it was found that the content of GA3 in different parts of the fruit was the highest,and it was several or even dozens of times of the other tested plant endogenous hormones.If the traditional R value[(ZT+GA3+IAA)/ABA]was used to comprehensively evaluate the effect of plant endogenous hormones on physiological fruit drop,the role of ZT and IAA in physiological fruit drop may be neglected.After the homogenization of the tested hormones,T value[(KZT+KGA3+KIAA)/KABA]was used for comprehensively analyzed,and it was found that the T value could be better reflect the relationship between endogenous hormones in different parts of fruit and physiological fruit drop.3.Analysis on the relationship between the early physiological fruit drop of W-murcott tangor and the content of four endogenous hormones in different parts of fruitThe W-murcott tangor with a strong facultative parthenocarpic ability was treated by pollination and non-pollination,meanwhile,the ovule and fruit development as well as physiological fruit drop process were observed.Plant endogenous hoemones were detected by HPLC in ovule,pulp,peel,whole fruit and fruit stalk at 25 DAF(25 day after flowering),32 DAF,39 DAF,50 DAF,60 DAF,90 DAF.The contents of endogenous hormones in the same part of W-murcott tangor was quite different in different developmental stages.32～39 DAF was the critical period of ovule development,the ovules showed obvious development or stagnation development.During this period,W-murcott tangor from the first physiological drops to the second physiological drops transition.39 DAF,the fruit of W-murcott tangor began to grow rapidly and enter the fruit swelling period.During 32～90 DAF,the peel of unpollinated W-murcott tangor has higher ZT content and that was the main existing part of ZT in fruit.During 25～50 DAF,the content of GA3 and IAA in the pulp of pollinated and unpollinated W-murcott tangor was higher,and the pulp was the main existing part of GA3 and IAA,while 50～90 DAF,peel became the main existing part of GA3 and IAA.During 25～50 DAF,the ABA content in the ovule was generally higher than that in other parts,and the ABA content in the ovule of the unpollinated W-murcott tangor is significantly higher than the pollinated W-murcott tangor.The abortive ovule may be the main existing of ABA in the fruit,which may be transported to the fruit stalk to induce the formation of the abscission layer and promote fruit drop.The T value analysis also showed that the pulp and peel had higher growth-promoting hormones content during 25-39 DAF,which were an important part to effect fruit development While peel was an important part to effect fruit development during 50～90 DAF.The growth and development of unpollinated W-murcott tangor were closely related to the content of endogenous hormones in pulp,peel and fruit stalk.During 25～60 DAF,the T value of the unpollinated W-murcott tangor whole fruit was higher than that of the pollinated W-murcott tangor whole fruit,which may be the result of more growth hormones produced by the pulp or peel of unpollinated W-murcott tangor.More growth hormones overcame the function of higher ABA produced by the abortive ovule and leaded to a reduced physiological fruit drop and guaranteed successful parthenocarpy of W-murcott tangor.4.Analysis on the relationship between pre-harvest fruit drops and content of four endogenous hormones in different parts of fruit in citrusFive citrus varieties with different pre-harvest fruit drop characteristics were selected as material,including Olinda Valencia orange,Seedless Valencia orange,W-Murcott tangor,Daidai and Shatianyou.Endogenous hormones in different parts of the fruit were detected at three time point after fruit physiological maturation on January 15,2018,January 27,2018,and March 4,2018.The content of endogenous hormones in different parts of varieties were quite different during pre-harvest fruit drop period,and the contents of ZT,GA3 and IAA in peel and fruit stalk were higher than other parts.The endogenous hormones contents in same parts of different varieties were quite different,and signal hormone content could not reflect its relationship with pre-harvest fruit drop accurately.ABA content was generally lower in the peel and fruit stalk of the varieties with lighter pre-harvest fruit drop,while ZT,GA3 and IAA showed no consistent performance.When compared with heavier pre-harvest fruit drop varieties,the T value in the peel and fruit stalk in lighter pre-harvest fruit drop varieties was obviously higher.The peel and fruit stalk may be more effective parts for analysis and judgement of the effects of endogenous hormones on pre-harvest fruit drop.5.Reconfirmation of the peel and fruit stalk as effective parts for analysis and judgement of pre-harvest physiological fruit dropDuring the pre-harvest fruit drop period,that is,December 10,2018 and December 25,2018,hybrids of TW cross breeding population characterized of different degree of pre-harvest fruit drop were selected as test materials.The endogenous hormones in the peel and fruit stalk were detected and analyzed by HPLC.The results showed that the T values could be better reflect the relationship between endogenous hormones and pre-harvest physiological fruit drop,when compared with the content of endogenous hormones and R value.The T values in the peel and fruit stalk of TW 201 and TW 140 with heavier pre-harvest fruit drop were significantly lower than that of TW 289 and TW 158 with lighter pre-harvest fruit drop.The results further verified that the peel and fruit stalk are the more effective parts for analysis and judgement of the effects of endogenous hormones on pre-harvest fruit drop.