【Objective】This study aimed to provide a theoretical basis for the strategic cultivation of plantations, and to increase the content of bioactive substances in leaves by investigating the effects of various nitrogen and phosphorus levels on the growth and bioactive substance content in Cyclocarya paliurus leaves.【Method】The research is conducted in the field and utilized a two-factor complete block design, incorporating three levels of nitrogen application (N0, N1, N2) and three levels of phosphorus application (P0, P1, P2), creating nine treatments with three replicates each. The growth indexes of seedlings and the contents of total flavonoids, total triterpenoids, total polyphenols and total polysaccharides in leaves were determined. A two-way ANOVA was used to compare the effects of nitrogen and phosphorus and their interaction on related indexes.【Result】The study found that different ratios of nitrogen and phosphorus significantly influenced both the growth and the bioactive substance content in C. paliurus leaves (P<0.05). The contents of total flavonoids, triterpenoids, and polyphenols in the leaves exhibited a parabolic seasonal variation, peaking in July, while the highest polysaccharides content was observed in September. Compared to the control treatment (N0P0), all nitrogen and phosphorus fertilizations enhanced tree growth, with the most substantial growth in tree height and ground diameter occurring in the N2P2 treatment. The effects of nitrogen and phosphorus ratios on the bioactive substances varied, displaying differences among the substances and the sampling months sampled. The variation in the contents of total flavonoid, total triterpenoids, total polyphenols and total polysaccharides in June ranged from 17.01 to 34.25 mg/g, from 39.11 to 55.77 mg/g, from 22.74 to 53.98 mg/g, and from 22.36 to 33.24 mg/g, respectively, while in October, the ranges were 16.95-35.26, 38.03-54.17, 19.78-33.98 and 23.33-32.46 mg/g, respectively. Correlation analysis revealed no significant relationship between the monthly growth in tree height and the content of bioactive substances (total flavonoid, total triterpenoids, total polyphenols and total polysaccharides) in the leaves (P>0.05). However, weakly negative correlations were observed between the ground diameter growth and the contents of total triterpenoids and total polyphenols in the leaves (P<0.05). Notably, there were significant positive correlations between the growth of tree height and ground diameter, as well as among the different bioactive substances (P<0.01).【 Conclusion】A high ratio of nitrogen and phosphorus addition benefit the growth of C. paliurus and enhances the content of total triterpenoids in the leaves. Sole nitrogen or phosphorus supplements boost the content of secondary metabolites, whereas a combined application tends to decrease the content of total polysaccharides. There is a noticeable trade-off between the growth of C. paliurus and the content of secondary metabolites in its leaves.
【Objective】Genomic simple repeat sequence (SSR) loci were analyzed by screening the whole genome of Cyclocarya paliurus. DNA molecular ID cards of 19 excellent medicinal clones of C. paliurus were constructed based on the newly-developed SSR primers. These genomic SSR markers could support further research, such as the evaluation of the germplasm resource, analysis of genetic diversity, and identification of cultivars/clones.【Method】The SSR loci were screened along with the whole genome of C. paliurus and were enriched and analyzed using MISA software. Subsequently, SSR primers were designed using Primer 3.0. Furthermore, a system for identifying clones of C. paliurus was constructed based on selected SSR markers with high reproducibility and stability.【Result】(1) We detected 89 741 SSR loci from the whole genome, with an occurrence frequency of 62.07%. (2) Among all SSR loci, the proportion of SSRs with a mononucleotide motif was the highest (62.67%) and a hexa-nucleotide repeat was the lowest (0.15%). Most of the repeated motifs in the SSR loci were dominated by (A/T)n. (3) The repeat number of mono-nucleotide and di-nucleotide motifs ranged from 6 to 16. With the increase in the repeat number, the frequencies of various SSR repetition types displayed a downward trend. (4) The length of the SSR sequences varied from 10 to 476 bp, and this length variation existed in different repetitive motifs. Additionally, the frequency of SSR occurrence tended to decrease as the repeat number increased. (5) We successfully designed 78 285 pairs of SSR primers using Primer 3.0. A total of 377 primer pairs were randomly synthesized for amplifying polymorphic SSR fragments, among which 75 pairs primers were successful. Moreover, quick response code DNA molecular ID cards for 19 medical-use clones of C. paliurus were constructed by five pairs of polymorphic SSR primers with a mono-nucleotide motif.【Conclusion】The frequency of genomic SSR loci was high, and there was variability in the type of SSR loci. Simple repeat sequences developed from the whole genome of C. paliurus could be effective candidate molecular markers with further applications in germplasm resource evaluation and fingerprint construction for multi-use clones of C. paliurus.
【Objective】To compare the differences in leaf phenotypes characters, anatomical structures and stomatal characteristics between diploid and tetraploid Cyclocarya paliurus, we obtained data on the leaf characteristics of C. paliurus with different ploidy.【Method】Diploid and tetraploid seedlings of C. paliurus were used as study materials, and 32 leaf traits were measured. Independent samples t-tests, a correlation analysis, and principal component analysis (PCA) were conducted.【Result】There were no significant differences between diploid and tetraploid seedlings for sawtooth width and the position of the maximum leaf width of the middle leaflet. For the other 30 leaf traits, there were significant or extremely significant differences between diploid and tetraploid seedlings. Among them, the number of leaflets of tetraploids was significantly less than that of diploids, while the area and specific leaf weight of compound leaves of tetraploids were significantly greater than those of diploids. The length, width and area of the terminal and middle leaflets of tetraploids were significantly greater than those of diploids, while the leaf shape index was significantly smaller than that of diploids. The sawtooth density of tetraploids was significantly smaller than that of diploids. The leaflet thickness, the upper epidermis thickness, lower epidermis thickness, palisade tissue thickness, and sponge tissue thickness of tetraploids were all significantly higher than those of diploids, while the ratio of palisade tissue to sponge tissue and cell tense ratio were significantly smaller than those of diploids. The stomatal length and width of tetraploids were significantly greater than those of diploids, while the stomatal density was significantly smaller than that of diploids. The correlation analysis results indicated that there were broad and significant correlations between the phenotypic characteristics, anatomical structure, and stomatal characteristics of C. paliurus. The indicators reflecting the differences of leaf characteristics between diploid and tetraploid C. paliurus were extracted by a PCA, including leaflet width, leaflet area, leaf shape index, sponge tissue thickness, stomatal length, palisade tissue thickness and cell tense ratio.【Conclusion】There were significant differences in leaf traits between diploid and tetraploid C. paliurus, and leaf phenotypic characteristics, such as leaflet width, leaflet area, and leaf shape index, can be used for the preliminary screening of diploid and tetraploid C. paliurus.
【Objective】To provide a theoretical basis for the introduction, selective breeding, and cultivation of Cyclocarya paliurus, the cold resistance of different families of C. paliurus under natural low temperature stress was evaluated.【Method】Using the current branches of C. paliurus from 23 families as materials, the relative electric conductivity (REC), malondialdehyde (MDA) content, peroxidase (POD) activity, superoxide dismutase (SOD) activity, soluble sugar (SS), soluble protein (SP), starch (ST) and free proline (Pro) content were determined after natural low temperature stress. A preliminary evaluation of the cold resistance of C. paliurus families was conducted through a principal component analysis (PCA) and cluster analysis.【Result】After natural low temperature stress, there were significant differences in the REC, MDA, POD and SOD activity, SS, SP, ST and Pro content among the current branches of the 23 families. The PCA found that the four principal components represented 72.4% of the information regarding the various physiological indicators. A cluster analysis showed that the cold resistance of 23 families could be divided into three categories based on the comprehensive score of the PCA for each family. The first category included only two families (SCMC31 and ZJTTS2) with the good cold resistance, and the compositive score ranging from 1.208 to 1.284. The second category indude 17 families (including GZSQ12, GXBS12, ZJFYS6, AHQLF8 and HBWF10) that exhibited moderate cold resistance, with composite scores ranging from -0.343 to 0.631. The third category included four families (GZSQ9, ZJTTS3, SCMC22, and SCMC30) with poor cold tolerance, with composite scores ranging from -1.259 to -0.745.【Conclusion】After natural low temperature stress, significant differences in the measured physiological indices were observed between the current branches of C. paliurus from different families (P < 0.05). Based on the PCA and cluster analysis results, there were different cold resistances among the 23 Cyclocarya paliurus families, which could be divided into three categories. The results not only provide a basis for an in-depth study of the cold tolerate mechanism of C. paliurus, but also for future screening of the cold tolerate genotypes of C. paliurus.
