Study On The Effect Of Raffinose Oligosaccharides On High Temperature Resistance Of Tomato Seedlings

Tomato is one of the main vegetable crops cultivated in China and the suitable temperature for plant growth are 13-33℃.The greenhouse temperature of summer facilities in China can reach above 35℃,even exceed 40℃.High temperature stress is one of the main limiting factors affecting tomato quality and yield.As a plant cell structural component,signal transduction and osmotic adjustment substance,raffinose family oligosaccharide plays an important role in plant resistance to stress.In order to study the role of raffinose family oligosaccharides in high temperature stress resistance of tomato,this test sprays galactinol(Gal),inositol(Ino),raffinose(Raf),sucrose(Suc)at the concentration of 3 mmol·L-1,galactose hydrolase inhibitor(DGJ)at the concentration of 5 μg·m L-1,and each half of the galactinol + galactose hydrolase inhibitor(G+D)by external application under high temperature stress,spraying the same amount of distilled water as a control(CK).The effects of raffinose on physiological indices of stress and the metabolism of raffinose in tomato under high temperature stress were studied.Inositol galactosidase,which is closely related to high temperature stress,was screened and gene overexpressing plants were constructed.The main results and conclusions are as follow:Plants sprayed with raffinose oligosaccharides were treated with high temperature stress at 38℃.After 6 days of stress treatment,the plants showed severe symptoms of wilting.Plants treated with G+D and Suc were slightly wilting.However,Gal,DGJ,Ino and Raf treatment significantly improved the high temperature resistance of the plants.Further analysis shows that RFOs pretreatment kept the relative conductivity,malondialdehyde and hydrogen peroxide content of tomato leaves at a low level,which reduced membrane damage,and increase the activity of SOD,POD and CAT in tomato leaves under high temperature.And reduced glutathione content,alleviating oxidative damage caused by high temperature stress.Analysis of the expression change of inositol galactosidase synthase(Gol S),raffinose synthase(RS),stachyose synthase(STAS)and α-galactose(AGAL)after exogenous spraying of RFOs under high temperature stress by q RT-PCR.The results showed that except for Sl RS4,other synthetase genes were up-regulated to a certain extent,but the expression level of the Sl Gol S gene family is closely related to high temperature stress.The expression of Sl Gol S gene family in tomato roots,stems and leaves under high temperature stress was further analyzed.The results showed that Sl Gol S4 in roots and stems did not respond to high temperature,and the expression of Sl Gol S gene family in tomato roots and stems was less,and it was expressed in leaves.There is a certain upward trend.Among them,the trend of Sl Gol S1 and Sl Gol S3 in leaves is more obvious than that of Sl Gol S2 and Sl Gol S4,and the expression level is more than three times.The prokaryotic expression vector was constructed and transformed into E.coli BL21 strain,and the positive strains of p ET32a(+)-Sl Gol S1,p ET32a(+)-Sl Gol S2,p ET32a(+)-Sl Gol S3 and p ET32a(+)-Sl Gol S4 recombinant plasmids were used as materials.After treatment at 37℃,45℃ and 50℃,the results showed that the colony growth was consistent after treatment at 37℃,and the colony decreased after 45℃ high temperature treatment,but the colony survival rate of p ET32a(+)-Sl Gol S1 and p ET32a(+)-Sl Gol S3 was higher than that of other,and the colonies were further reduced after treatment at 50℃,but the p ET32a(+)-Sl Gol S1,p ET32a(+)-Sl Gol S3 colonies survived.The rate is still high,indicating that Sl Gol S1 and Sl Gol S3 respond to high temperature and can speculate the high temperature tolerance of E.coli.The recombinant plasmid of Sl Gol S1 and Sl Gol S3 was transformed into Agrobacterium tumefaciens,Agrobacterium tumefaciens-mediated infecting tomato cotyledons,and the callus was cultured and the tomato seedlings were obtained.The positive plants were identified by PCR amplification,and 3 strains of Sl Gol S1 overexpressing T0 plants and 2 strains of Sl Gol S3 overexpressing T0 plants were obtained,which provided important materials for further study on tomato high temperature resistance.