| Paeonia lactiflora Pall.has been known and cultivated for nearly 4,000 years during the whole Chinese civilization history, which is mainly distributed or cultivated in north, northwest and northeast regions of China. However, landscape utilization of P. lactiflora is rather rare in south region of the Yangzi River, for instance, Zhejiang Province. Just like the Northward Plantation of Prunus mume Sieb., the Southward Plantation of P. lactiflora is also a classic problem of landscape plant, ornamental horticulture and landscape architecture, which have been paid much attention in theory for many years, but lacked of scientific practice. A total of 30 P. lactiflora cultivars introduced from Heze of Shandong and Pan’an of Zhejiang were selected as materials in this research. Our studies focusing on P. lactiflora for four years covered landscape use status in Hangzhou City, observation for introduction and cultivation, physiological and biochemical response to high temperature stress, heat resistance evaluation, comprehensive evaluation for growth and development, chilling requirement for the release of underground renewal bud dormancy, flowering regulation, transcriptomic sequencing, research on differentially expressed gene related to bud dormancy, and crossbreeding between the north and south peonies. We hope these researches could benefit for the Southward Plantation of P. lactiflora to a certain degree.1 Landscape use status in Hangzhou and observation for introduction and cultivationCompared with Paeonia suffruticosa Andr., it is rather scanty that the used cultivars and arrangement techniques of P. lactiflora in Hangzhou park green space. P. lactiflora cultivars are planted intensively in Herbaceous Peony Garden of Fish Viewing at the Flower Pond, however, their growth and flowering degraded seriously year after year. The main causes of these phenomena are as followed:(1) the high temperature in winter of Jiangnan than North Area leads to insufficient chilling accumulation and abnormal growth and flowering; (2) the high temperature in spring brings about excessive growth and drooping of flower stem, and also results in too short flowering time; (3) the muggy climate in summer gives rise to serious sunburn, heat, pest and disease damage. Breeding new cultivars with low chilling requirement, simple flower type, strong flower stem and high heat resistance is very important for the Southward Plantation of P. lactiflora. Scientific research, new cultivar spreading and industrialized production could also contribute to achieve the Southward Plantation of P. lactiflora.The first step is resource introduction for realizing the Southward Plantation of P. lactiflora. We introduced 29 ornamental cultivars from Heze of Shandong and a medicinal cultivar named’Hangbaishao’from Pan’an of Zhejiang, and carried out observation and measurement for growth and development for four years. The data were collected during the periods of sprouting, growth, flowering and withering. P. lactiflora’Hangbaishao’, ’Guanyinzuo’, and’Qihua Shuanglu’grew vigorously and bloomed exuberantly. These cultivars are suitable for widely use in Hangzhou. On the other hand,’Hongjinqiu’,’Qiaoling’ and ’Wuhua Longyu’ showed up the worst growth and flowering.’Hangbaishao’should be a cultivar group with various flower colors, but its classification is not very clear until now, so we studied it as a cultivar in this research.2 Physiological and biochemical responses to the high temperature, evaluation for heat resistance and growth adaptabilityAccording to the heat resistance evaluation for 30 P. lactiflora cultivars based on the heat injury index (HII), relative chlorophyll content (RCC) and semilethal temperature (LT50), we found ’Hangbaishao’,’Yangfei Chuyu’and’Hongjin’gang’had the strongest heat resistance, whereas’Qiaoling’,’Dafugui’and’Qingwen’had the lowest resistance. Compared with the cultivars with lower heat resistance, the cultivars with stronger heat resistance had lower relative electrical conductivity (REC) and malondialdehyde content (MDA), and higher relative water content (RWC), absolute chlorophyll content (CC), soluble sugar content (SSC), soluble protein content (SPC), ascorbate peroxidase activity (APX), and values of Fm, Fv/Fm,ΦPSⅡ, qp and ETR. Superoxide dismutase activity (SOD) changed unsteadily.Two methods, which abbreviated as SF and SF+PCA method in this research, were used to evaluate the heat resistance of ten representative cultivars based on the physiological and biochemical data. The full name of SF is subordination function method, and SF+PCA is the abbreviation of the method of subordination function in combination with principal component analysis. The results indicated that ’Hangbaishao’ and ’Yangfei Chuyu’ had the highest heat resistance, while’Qingwen’and’Dafugui’had the lowest resistance. Based on the analytical hierarchy process (AHP),’Qihua Shuanglu’,’Hangbaishao’and ’Shanhehong’had the best comprehensive growth adaptability, all of which are suitable for landscape use in Hangzhou and other south regions of the Yangzi River, or preferentially use as parents in crossbreeding for excellent cultivar.’Qiaoling’,’Qingwen’and’Hongjinqiu’showed the worst comprehensive adaptability.3 Chilling requirement, flowering regulation, and dormancy physiology and biochemistry of ’Hangbaishao’under the natural and aitifical low temperature’Hangbaishao’needed at least 540 cumulative chilling hour (CH) and 665 cumulative chilling unit (CU) to break its underground renewal bud dormancy. A total of 795 CH and 906 CU is optimum to meet the chilling accumulation for bud dormancy release. Under the artificial chilling treatment,’Hangbaishao’needed 672 CH and 590.69 CU to break bud dormancy, and grew and bloomed vigorously.’Hangbaishao’could bloom 35 to 40 days in advance under the natural chilling combined with greenhouse cultivation. The forcing of sprouting and flowering was limited under 5-azacytidine treatment. Using GA3 could force sprouting obviously, but also led to inevitable plant lodging and withering. Under the artifical chilling,’Hangbaishao’could bloom 60 days in advance after four weeks treatment in refrigeration storage (0-4℃) and 300 mg/L humic acid (HA) treatment.During the bud dormancy under the natural low temperature, IAA/ABA fluctuated, starch content dropped, while SSC and peroxidase (POD) activity rised steadily; when the bud sprouted, SSC and POD activity had a sharp decline, starch content changed unobviously and always showed the low level.5 Transcriptomic sequencing of the’Hangbaishao’underground renewal budsThe bud dormancy of’Hangbaishao’could be divided to five stages:endodormancy, transition from endodormancy to ecodormancy, ecodormancy, ecodormancy release, and bud elongation and sprouting. We performed de novo transcriptome sequencing using the Illumina RNA sequencing platform for the underground renewal buds of P. lactiflora’Hangbaishao’, to study the molecular mechanism underlying its bud dormancy transition (the period from endodormancy to ecodormancy) and release (the period from ecodormancy to bud elongation and sprouting). A total of 207,827 (mean length= 828 bp) and 51,481 (mean length= 1250 bp) unigenes were assembled by using two assembly methods named "Trinity" and "Trinity+PRICE", respectively. Based on the data obtained by the latter method,2,057 differentially expressed genes (DEGs) were screened out. Annotation of DEGs were also studied, for instance, GO functional enrichment, expression profile, clustering and comparison of DEG number between different dormancy stages, etc..6 Expression of differential genes in’Hangbaishao’buds during the different dormancy stagesDEGs could be divided into three categories:the DEGs related to environmental response, metabolism, and cell growth and development. During the bud dormancy, expression of suppresspor of overexpression of constansl (SOC1) and late elongated hypocotyl genes showed first increase and then decrease, which might be related to chilling requirement fulfillment, cold resistance and circadian rhythm. Phytochrome (PHY), heat shock protein (HSP) and osmotin (OSM) genes up-regulated during sprouting, which might be induced by the change of light and temperature, and contributed to break the bud dormancy and stem elongation. The genes of aquaporin (AQP), dehydrin (DHN), β-amylase, H+-ATPase, auxin-repressed protein (ARP), repressor of GA (RGA), GA20 oxidase (GA20ox), ethylene responsive element binding factor (ERF) and peroxidase (PER) might be associated with material transportation and metablisms of water, carbohydrate, energy, phytohormone and antioxidative enzyme. All these DEGs expressed actively, might promote or inhibit the bud dormancy release. The genes of cyclin (CYC)、expansin (EXP). histone and regulator of chromosome condensation 1 (RCC1) are important for cell cycle and division, most of which up-regulated when the bud sprouting and elongation. The expression profiles of DEGs were mainly based on the value of fragments per kilobase of exon model per million mapped reads (FPKM value) of unigenes obtained from transcriptomic sequencing. It is necessary to adopt other molecular biology methods to confirm these results and excavate more important DEGs closely related to bud dormancy.7 Crossbreeding between the north and south cultivarsThe pollen viability of ’Hangbaishao’ was 55.60% based on four measurement methods, which were triphenyl tetrazolium chloride (TTC), aceto carmine, I2-KI staining and germination method. The pollen viability of ’Hangbaishao’was high enough for crossbreeding. The seeds could not germinate which obtained from crossbreeding between different Heze cultivars. But when ’Hangbaishao’ acted as the male parent, all of hybrid seeds could germinate and seedlings could grow vigorously, thus it can be seen that it is possible to produce excellent cultivars with good adaptability to Zhejiang climate environment by crossbreeding between the north and south herbaceous peony resources... |
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