| Paeonia ostii is one kind of medicinal peony types originating in the Fenghuang Mountain area of Tongling in Anhui Province(Its root bark (Cortex Moutan) is known as famous genuine medicinal materials) and it also has high ornamental value. P. ostii is always used as an excellent rootstock for ornamental peony cultivars, because of its strong root system and strong stress tolerance. Predecessors and our study found that P. ostii grew well in Fenghuang Mountain of Tongling in Anhui where was an typical area of copper mine tailing, and Cortex Moutan is famous for its high yield and good quality in this area. It indicate that P. ostii has a good ability to adapt to the copper contaminated soil. In order to study the mechanism of copper tolerance in P. ostii., We investigated growth and physiological responses of P. ostii under copper stress to learn its physiological mechanism. Furthermore, we analysised the changes of P. ostii at the level of transcriptome under copper stress to look for copper stress response genes and learn the molecular mechanism underline. It could provide scientific evidences for the exploitation and remediation of copper contaminated soil using P. ostii. In view of P. ostii is used as rootstock for ornamental peony, a further experiment was conducted to study if the grafting ornamental peonies had the samilar copper tolerance like P. ostii, which could provide theoretical basis for broaden the scope of P. suffruticosa planting and trying to plant P. suffruticosa in copper contaminated soil to reduce copper content and beautify the environment1. Effects of copper stress on growth and physiological characteristics of P. ostii.Copper tolerance in P. ostii under different concentrations of Cu treatments (0,300mg·kg-1,600mg·kg-1,900mg·kg-1and1200mg·kg-1) were studied by soil culture method. The results showed that there were not obviously growth inhibition in P. ostii under Cu stress; The concentration of copper (300mg·kg-1) can promote the increase in plant height and the length of the new branches; The concentration of copper (600mg·kg-1) can promote root collar thickening and taproot elongation; The low concentration of copper (≤600 mg·kg-1) had no significant effect on the ratio of chlorophyll a/b and chlorophyll content; The activity of SOD can be induced when1200mg·kg-1for Cu was supplied. The activity of SOD in root and POD activity in leaves and roots under copper stress increase firstly and then decreased. The content of MDA was changed significantly only at Cu1200mg·kg-1treatment. The content of Ca and Mn was not affected under copper stress, the content of K in leaves was significantly decreased under600mg·kg-1and900mg·kg-1Cu treatment, but the content of Mg in leaves increased under600mg·kg-1Cu treatment, the absorption of Zn was inhibited when the concentration of copper was under600mg·kg-1or even more, while the absorption of Fe of the root was enhanced under Cu treatment, but the content of Cu in leaves increased significantly under copper stress. The content of P in leaves and stems increased significantly under Cu treatment (900and1200mg·kg-1), while the absorption of P in roots began to increase significantly when the concentration of Cu was600mg·kg-1.2. Antioxidase activity and other physiological characteristics of P. suffruticosa.Growth, physiology, and element absorption of P. suffruticosa (taking P. ostii as the root stock) under different concentrations of Cu treatments (low,300mg·kg-1; middle,600mg·kg-1; and high,900mg·kg-1) were studied by soil culture method. The results showed that most of the morphological indices were not obviously affected by300mg·kg-1and600mg·kg-1Cu treatments, and600mg·kg-1Cu treatment can promote rhizome diameter. Under all copper treatments, chlorophyll content had no significant changes, but the ratio of chlorophyll a and b was decreased under600and900mg·kg-1Cu treatments. The activity of SOD can be induced by Cu treatment, but it was less affected by900mg·kg-1. The activity of POD increased significantly under300mg·kg-1Cu treatment, while inhibited by600mg·kg-1and900mg·kg-1Cu treatment. The content of MDA was not changed significantly under low Cu treatment, but it began to increase under middle and high Cu treatments. The root accumulated most of the Cu ions through the retention effect, and the Cu content of shoots was maintained at low levels. The content of Zn in leaves decreased significantly under all Cu treatments, while it was inhibited only by high Cu treatment in roots. In addition, all Cu treatments have not led to a significant reduction in the P, K, Ca, Mg, Fe and Mn absorption.3. The transcriptome analyses of P. ostii.In this study we used hydroponic culture method, and P. ostii. were exposed to100μM CuSO4for stress experiments. Total RNAs were isolated from root, stem and leaf atl,6, 12and24h of copper stress. Each of sample RNA from CK and copper treatment was pooled (in equal amounts) respectively to obtain two RNA samples. Then, we performed transcriptome sequencing of CK library and copper library via RNA-Seq. In the CK library, there are53221924clean reads, which contain4789973160nucleotide sequences.160071contigs were obtained of which the average length is300bp, the total length is4789973160bp and N50was496bp.82302unigene were matched out of which the average length is600bp, the total length is49382245bp and N50was985bp. In the copper library, there are52068126clean reads, which contain4686131340nucleotide sequences.144049Contigs were obtained of which the average length is307bp, the total length is44238621bp and N50was506bp.82302unigene were matched out of which the average length is600bp, the total length is49382245bp and N50was929bp. The Unigenes functional annotations show that45684Unigenes were matched to the Nr database and28821Unigenes were matched to the Swiss-Prot database. There are4324differentially expressed genes in copper stress, the number of up-regulated gene and down-regulated gene were2207,2117respectively; KEGG pathway enrichment analysis of differentially expressed genes show that there are118pathway enriched. The genes which significantly differentially expressed in these pathway may play an important role in the regulation of P. ostii response to copper stress.Under the copper stress, expression of genes related to multicopper oxidase and metallothioneins were up-regulated, however some members of the ZIP and ABC gene family were down-regulated. The HSPs gene family in particular small molecule heat shock protein are also involved in the regulation of P. ostii response to copper stress, the expression of which had high expression. |
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