[Transcriptome analysis of rhizome of Polygonatum cyrtonema and identification of candidate genes involved in biosynthetic pathway of steroidal saponin].

To enrich the transcriptome data in rhizome of Polygonatum cyrtonema seedlings, identify candidate functional genes involved in steroidal saponin biosynthesis and provide genetic resources for the research on anabolism pathway and regulatory mechanism of active components in P. cyrtonema, Illumina platform was applied to perform transcriptomic sequencing of rhizome of P. cyrtonema, followed by a series of bioinformatics analysis on RNA-seq data, including de novo assembly, annotation, classification and metabolic pathway analysis of the assembled unigene. Meanwhile, a deep analysis on the steroidal saponin biosynthesis in secondary metabolism pathway was performed. The results showed a total of 126 546 unigene were obtained by de novo transcriptome assembly, of which 47 226 were annotated. Of these, 16 499 unigene were mapped to 132 specific pathways, of which 2 768 were identified to be involved in 22 secondary metabolic pathways. One hundred and thirteen unigene were identified from the transcriptome database, which encoded 27 metabolic enzymes associated with steroidal saponin biosynthesis and shared similarity with 45 functional genes in Arabidopsis thaliana. In conclusion, a series of candidate functional genes, which might be involved in steroidal saponin biosynthesis, were selected from the transcriptome database of P. cyrtonema rhizome. Further investigation of these candidate genes will provide insight into their actual functions in the steroidal saponin biosynthetic pathway in P. cyrtonema. In addition, this study also provide abunant reference data for transcriptome characterization of P. cyrtonema and has important significance for functional genomics of P. cyrtonema.

[Selection and validation of internal reference genes for qPCR in Polygonatum cyrtonema tubers at different development stages and in response to abiotic stress].

In order to analyze the expression of genes involved in steroidal saponin biosynthesis pathway in Polygonatum cyrtonema tubers, it is very important to select internal reference genes that are stably expressed at different development stages and in response to abiotic stress. According to the previously established P. cyrtonema transcriptome database and reported internal reference genes in plant, this study systematically analyzed eight candidate internal reference genes including histone H2 A, glyceraldehyde-3-phosphate dehydrogenase, ACTIN, ß-tubulin, ubiquitin-conjugating enzyme-E2-10, elongation factor 1-alpha isoform, 18 S rRNA and α-tubulin 4 for expression stability in P. cyrtonema tubers at different development stages and in response to methyl jasmonate(MeJA) stress by using Real time fluorescence quantitative PCR(qPCR). Based on the statistical analysis of qPCR results by using GeNorm, NormFinder and BestKeeper softwares, the expression of ubiquitin-conjugating enzyme-E2-10 and elongation factor 1-alpha isoform are the most stable in P. cyrtonema tubes at different development stages and in response to MeJA stress. The two internal reference genes were further validated by analyzing the expression of 4 genes involved in steroidal saponin biosynthesis pathways. In conclusion, ubiquitin-conjugating enzyme-E2-10 and elongation factor 1-alpha isoform can be used as the most appropriate internal reference genes for qPCR analysis in P. cyrtonema. This study also provide a foundation for future investigate the molecular mechanism of steroidal saponin biosynthesis pathways in P. cyrtonema.

Effects of 60-day NO2 fumigation on growth, oxidative stress and antioxidative response in Cinnamomum camphora seedlings.

OBJECTIVE: To study the oxidative stress and antioxidative response of Cinnamomum camphora seedlings exposed to nitrogen dioxide (NO(2)) fumigation. METHODS: Measurements were made up of the growth, chlorophyll content, chlorophyll fluorescence, antioxidant system and lipid peroxidation of one-year-old C. camphora seedlings exposed to NO(2) (0.1, 0.5, and 4 microl/L) fumigation in open top chambers over a period of 60 d. RESULTS: After the first 30 d, 0.5 and 4.0 microl/L NO(2) showed insignificant effects on the growth of C. camphora seedlings. However, exposure to 0.5 and 4.0 microl/L NO(2) for 15 d significantly reduced their chlorophyll content (P<0.05), enhanced their malondialdehyde (MDA) content and superoxide dismutase (SOD) activity (P<0.05), and also significantly reduced the maximal quantum yield of PSII in the dark [the ratio of variable fluorescence to maximal fluorescence (F(v)/F(m))] (P<0.05). In the latter 30 d, 0.5 microl/L NO(2) showed a positive effect on the vitality of the seedlings, which was reflected by a recovery in the ratio of F(v)/F(m) and chlorophyll content, and obviously enhanced growth, SOD activity, ascorbate (AsA) content and glutathione reductase (GR) activity (P<0.05); 4.0 microl/L NO(2) then showed a negative effect, indicated by significant reductions in chlorophyll content and the ratio of F(v)/F(m), and inhibited growth (P<0.05). CONCLUSION: The results suggest adaptation of C. camphora seedlings to 60-d exposure to 0.1 and 0.5 microl/L NO(2), but not to 60-d exposure to 4.0 microl/L NO(2). C. camphora seedlings may protect themselves from injury by strengthening their antioxidant system in response to NO(2)-induced oxidative stress.

[Effects of NO2 on Cinnamomum camphora seedlings growth and photosynthesis].

A 2-month fumigation experiment was conducted with opened top chambers to study the effects of different concentration (0.1, 0.5, and 4.0 microl x L(-1)) NO2 on the growth and photosynthesis of 1-year Cinnamomum camphora seedlings. Fumigation with 0.1 and 0.5 microl NO2 x L(-1) promoted the growth of the seedlings, while with 4.0 microl NO2 x L(-1) was in adverse. The diurnal variation of net photosynthetic rate (P(n)) presented two-peaks, with an obvious depression in midday. 0.5 microl NO2 x L(-1) increased the P(n), the maximum of P(n) reached 8.542 micromol CO2 x m(-2) s(-1); 4.0 microl NO2 x L(-1) decreased the P(n) in most period of time; while the effect of 0.1 microl NO2 x L(-1) varied with time period. Fumigation with 0.5 and 4.0 microl NO2 x L(-1) increased the maximal and minimal values of stomatal conductance (G(s)) and intercellular CO2 concentration (C( i)), while 0.1 microl NO2 x L(-1) increased the maximal and minimal values of C(i) but decreased the maximal and minimal values of G(s). At the middle and late stages of fumigation, the mean P(n) of the seedlings treated with 0.5 microl NO2 x L(-1) was significantly higher than that treated with 0.1 and 4 microl NO2 x L(-1). At the early stage of fumigation, 0.5 and 4.0 micro NO2 x L(-1) significantly decreased the maximal PS II efficiency (F(v)/F(m)); and at the late stage, 4.0 microl NO2 x L(-1) still decreased the F(v)/F(m) significantly.