| Heavy metals are a kind of serious pollutants in soil and water,with the development of industry,the stress of heavy metal pollution on plant growth is increasing.It seriously endangers the quality of soil environment and restricts the normal life activities of plants.Cadmium(Cd),as one of the most toxic heavy metal elements,can cause toxic reactions in plants and animals at low concentrations.Moreover,as heavy metal Cd is a cumulative toxin,its toxicity and harm to human body will expand after it enters the human body through the food chain and becomes enriched.In addition,its half-life is long and difficult to be discharged to the body,which will continue to cause harm to the human body,so the treatment of heavy metal Cd pollution is urgent.Phytoremediation is an environmentally friendly technology for the treatment of heavy metals,but there are still few effective remediation plants applied to the treatment of Cd.Therefore,it is of great application value to study the mechanism of Cd resistance in plants and improve their Cd resistance.In this paper,we took Populus ussuriensis as the research object,and preliminatively investigated the function of PuERF9 transcription factors under Cd stress.The main results were as follows:(1)The expression pattern of PuERF9 under 200 μM ABA,6%PEG6000,200 mM NaCl,100 μM CdCl2,1.2 mM ZnSO4 and 10 μM low phosphorus were analyzed by qRT-PCR,the results indicated that PuERF9 could be significantly induced by 100 μM CdCl2 in both roots and leaves of P.ussuriensis.We cloned the proPuERF9 from P.ussuriensis,and the proPuERF9::GUS transgenic P.ussuriensis was obtained by Agrobacterium-mediated stable transformation.It was found that GUS histochemical staining was enhanced in root tips and leaves of transgenic plants treated with CdCl2 stress.PlantCare was used to predict the PuERF9 gene promoter elements,and we found that there were MeJA,ABA response elements,cis-acting regulatory element essential for the anaerobic induction and cis-acting element involved in defense and stress responsiveness.(2)The subcellular localization of transcription factor PuERF9 was conducted by the transient protoplast transformation method of Populus alba × P.Berolinensis callus,and it was found that it was located in the nucleus.The transcriptional activity of PuERF9 was analyzed by yeast autoactivation assay and transient expression of protoplast,and it was determined that PuERF9 had no transcriptional activation activity and was a transcriptional repressor.(3)We constructed the overexpression vector(PuERF9-OE)and repression vector(PuERF9-RNAi)of PuERF9,and the genetic transformation of P.ussuriensis was carried out by A.tumefaciens-mediated leaf disc method.At the DNA level,the transgenic strains of poplar were detected and verified by RT-qPCR.Wild type and transgenic tissue culture of P.ussuriensis seedlings were treated with 100 μM CdCl2 to compare their cadmium resistance.The results showed that the PuERF9-OE strain showed better growth than the wild type in the medium supplemented with 100 μM CdCl2,but the PuERF9-RNAi strain showed weaker growth than the wild type.The physiological indexes of each strain were determined and analyzed,we found that overexpression of PuERF9 gene can reduce the damage of plant cells under Cd stress,reduce the accumulation of ROS,increase the contents of proline and chlorophyll in plants.These results suggest that PuERF9,as a transcription repressor localized in the nucleus,may participate in the process of response to cadmium stress of P.ussuriensis sinense and enhance the cadmium resistance of populus sinense by removing reactive oxygen species. |
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