Studies On Cold Resistance Of Sugar Transporter DlSWEET1 Gene In Longan

Longan(Dimocarpus longan Lour.),as a typical subtropical evergreen economic fruit tree in China,is sensitive to temperature and has the characteristics of liking temperature and avoiding freezing and poor cold resistance.Low temperature is an important factor restricting the growth and development of longan fruit trees.Therefore,the study on the changes of physiological indexes of longan under low temperature stress will help to take countermeasures in real production and reduce the harm caused by low temperature.SWEET protein is a sugar transporter containing seven transmembrane helices and two MtN3 conserved domains,which is widely found in plants,animals and prokaryotes.It has been suggested that SWEETs sugar transporter plays an important role in plant growth,reproductive development,biological and abiotic stress.In this study,the changes of longan leaf physiological indexes under low temperature stress were first determined,the whole genome of the longan SWEET gene family was further identified and analyzed by the’Honghezi’ longan genome data,and the response response of DlSWEET genes to abiotic stress(low temperature,high temperature,drought and salt)was studied by fluorescence quantification technology,and DlSWEET1 was screened out to play an important role in the cold resistance of longan The functional study of DlSWEET1 gene was carried out by transforming Arabidopsis thaliana and combining physiological and molecular biology,and the genetic transformation of longan embryonic callus was carried out by gene gun method,laying a foundation for further study of the function of DlSWEET1 gene.The main findings are as follows:1.Physiological changes of longan leaves at low temperatures:With the extension of treatment time,the MDA content of leaves rises first and then falls and then rises;POD activity also shows a trend of rising first and then falling and then rising;SOD activity decreases and then rises;the content of total chlorophyll,chlorophyll a,chlorophyll b generally shows a trend of first falling and then rising;soluble protein and soluble sugar content show a trend of first increasing and then decreasing,showing adaptability to low temperature environment.2.Longan SWEET gene family analysis:According to the published longan genome information,20 longan SWEET family members were identified based on the amino acid sequence of Arabidopsis SWEET protein as the query sequence,and renamed to DlSWEET1-DlSWEET16b according to their phylogenetic relationship with Arabidopsis thaliana SWEET gene.Analysis of tissue-specific expression patterns suggests that the DISWEET gene may play a variety of roles in the development of longan tissue.qRT-PCR analysis showed that six DlSWEETs(DlSWEET1/2a/3a/10c/16a/16b)may be involved in the non-biological stress response of the fruit,of which only the expression of the DlSWEET1 gene was upregulated under cold stress,predicting that it may play an important role in the anti-cold response of longan.3.Study on the function of DlSWEET1 gene:Constructed an overexpression vector of DlSWEET1 gene,obtained transgenic Arabidopsis strains using inflorescence infestation as material,and performed phenotypic observation and abiotic stress experiments.The results showed that the overexpression of DISWEET1 gene did not significantly affect the growth and development process of Arabidopsis thaliana plants.In low temperature stress,by observing the growth and survival status of plants and measuring the POD activity and soluble sugar content of plants,it was proved that DlSWEET1 gene can strengthen the cold resistance of Arabidopsis thaliana.4.Establishment of genetic transformation system mediated by genetic marksmanship of longan callus:Based on the direct somatic embryogenesis system of longan,an effective genetic transformation system was initially established by adjusting the antibiotic concentration,pre-infiltration treatment time,post-infiltration treatment time,vacuum,range and bombardment distance,and the number of bombardments,with the instantaneous transformation efficiency of GUS as the index.