Transcriptome Analysis Of Reaumuria Trigyna Under Cadmium Stress And Functional Vadiation Of A Cadmium Transporter RtCAX3

Environmental stress such as salt,cadmium seriously restrict the growth and yield of plants.It is of great significance for soil bioremediation and breeding of stress-resistant varieties to utilize the stress-resistance species.Reaumuria trigyna,a small shrub belongs to the genus Reaumuria L.in the family Tamaricaceae,it has strong adaptability to adverse environment.Its mechanism of salt tolerance and drought resistance has been studied in depth,but its ability to resist heavy metal stress has not been paid more attention,and the underlying mechanism is still unclear.In this study,we performed transcriptome sequencing of R.trigyna seedlings under cadmium stress.Then,a cadmium transporter gene RtCAX3 was cloned,and its function were analyzed in transgenic Arabidopsis.This study laid a solid foundation for exploring the resistance mechanisms of R.trigyna in response to heavy metal stress,and it also provides a theoretical basis for using this species to ameliorate heavy metal contaminated soil.The main results are as follows:1.In this study,RNA-Seq technology was used for transcriptome analysis of R.trigyna seedlings under cadmium stress.A total of 115377 Unigenes were obtained,and 45485 Unigenes were annotated into 7 databases including NR and GO.GO analysis showed that the number of Unigenes enriched in"Cellular process"items was the largest.COG analysis showed that Unigenes were enriched in"Post-translational modification,protein turnover,chapterone"items.2.Analysis of cadmium transporters was conducted in the cadmium-stress transcriptome of R.trigyna.The results showed that NRAMP,HMA,MTP,CAX,ZIP,YSL and ABC protein families that mediate cadmium transport were expressed in both the control and the Cd treatment groups,and 9 genes were differentially expressed.Among the 9 DEGs,1 NARMP gene was down-regulated while the other 8 genes（1 CAX gene,5 ZIP genes and 2 YSL genes）were up-regulated.3.RtCAX3 was screened and cloned base on the cadmium-stress transcriptome data of R.trigyna.The ORF of RtCAX3 gene was 1326 bp,encoding a protein composed of 441 amino acid.RtCAX3 gene was expressed in roots,stems and leaves of R.trigyna.It could be induced by salt,drought,Cd Cl₂and low temperature treatments.Subcellular localization result showed that it was localized in the vacuolar membrane.4.We transformed RtCAX3 gene into Arabidopsis to analyze its function.The wild type（WT）and transgenic plant were treated with salt or Cd Cl₂stress.The growth condition,antioxidant enzyme activity and proline content of transgenic seedling were significantly higher than those of WT,while the malondialdehyde and hydrogen peroxide content is significantly lower than those of wild type.Six stress-responsive genes were induced in transgenic Arabidopsis and their expression levels were significantly higher than those of wild type.Therefore,RtCAX3 may improve the salt and cadmium tolerance of transgenic Arabidopsis by enhancing the activity of antioxidant enzymes,maintaining the osmotic balance and promoting the expression of stress-responsive genes.