Resistance Identification Of Genes TaVDAC1 And TaRS In Transgenic Arabidopsis

Voltage-dependent anion channels,VDAC,of eukaryotic mitochondrial outer membranes play key role in the process of plant growth,apoptosis and stress.The Raffinose family oligosaccharides,RFOS,in higher plants not only play an important role in seed germination and maturation,but also associated with reducing the effects of environmental stress,including oxidation,extreme temperatures,salinization and drought.Our previous studies in our laboratory,the genes TaVDAC1 and Ta RS were cloned from wheat,moreover,analysis of bioinformatics,subcellular localization,and expression patterns of two genes has been done.Further,the genes were transferred into Arabidopsis and the T3 transgenic lines seeds were harvested.Based on which,the main content of this study is to identify the resistance of TaVDAC1 and Ta RS transgenic Arabidopsis thaliana,the results obtained are as follows:1.DNA positive identification of TaVDAC1 transgenic Arabidopsis thaliana T3 lines were carried out,and RNA were extracted from TaVDAC1 positive lines,further,overexpressed lines OE-3,OE-4,OE-6 were screened by q RT-PCR.2.The germination rate was statistically analyzed under Na Cl and mannitol stress conditions,the results showed that the germination rate of TaVDAC1 transgenic lines were significantly higher than that of WT,conversely,it showed sensitivity to ABA treatment;The green rates of cotyledon under salt and drought treatment were analyzed,it was found that the green rate of TaVDAC1 transgenic lines were significantly higher than WT;Under the stress treatment of Na Cl and mannitol,the scanning results of root length showed that the number of lateral roots and total root length of TaVDAC1 overexpressing lines were significantly higher than that of WT,Under ABA treatment,the main root growth of TaVDAC1 transgenic lines was greatly inhibited;In addition,the leaves of transgenic lines showed a yellowing phenotype,combined with the sensitive phenotype of germination and root growth,indicating that TaVDAC1 may participate in ABA signaling pathway;NBT staining demonstrated that the overexpressing lines had lower levels of active oxygen in the body under salt treatment;The water loss of rosette leaves were measured in vitro,and the results showed that the water loss rate of TaVDAC1 overexpressing lines were lower than that of WT;After long-term natural drought,transgenic lines were observed to be stronger than wild type;After low temperature treatment,the survival rate of the transgenic lines was higher than that of the wild type.It is shown that TaVDAC1 enhances the tolerance of Arabidopsis to salt,drought,low temperature,but enhances the sensitivity to ABA.3.The effect of TaVDAC1 on the expression of some stress-related genes(such as RAB18,WRKY28,P5CS1,NCED3,RD29 A,COR47,KIN1,COR15 A,etc.)were analyzed.The results showed that stress-related genes in WT and TaVDAC1 overexpressing lines were significantly up-regulated in response to salt,drought and cold stress,however,the transcriptional level of ABA-related stress response genes in overexpressing plants were generally lower than that of WT.4.DNA positive identification of Ta RS transgenic Arabidopsis thaliana T3 lines were carried out,and RNA were extracted from Ta RS positive lines,further,overexpressing lines L5-3,L6-4,L7-2 were screened by q RT-PCR.5.Under Na Cl and mannitol treatment,the germination rate was counted and the results showed that the germination rate of Ta RS overexpressing Arabidopsis was significantly lower than that of WT.The growth of roots under salt and drought stress was observed,the results showed that there was no difference between Ta RS overexpressing lines and WT,but the fresh weight of overexpressing lines was significantly smaller than WT.Which indicated that Ta RS overexpressing Arabidopsis is more sensitive to salt and drought.The results of this study will provide an experimental basis for elucidating the resistance function of TaVDAC1 and Ta RS and its mechanism of stress response.