Effects of Drought Stress Simulated by Polyethylene Glycol on Seed Germination, Root and Seedling Growth, and Seedling Antioxidant Characteristics in Job’s Tears

Two Job’s tears cultivars, yy18-1 (high resistance to drought stress) and yy12-7 (susceptible to drought stress) were used to investigate the responses of seed germination, root and seedling growth, and seedling antioxidant characteristics to drought stress simulated by polyethylene glycol (PEG) 6000 solutions with 0, -0.05, -0.1, -0.15, and -0.2 MPa osmotic potentials. The results showed that the germination energy, germination rate, germination index, root and seedling lengths, root and seedling diameters, root and seedling fresh masses, root and seedling dry masses, and seedling relative water content (RWC) decreased with the decrease of the osmotic potential of PEG 6000 solution. The contents of hydrogen peroxide (H2O2), malondialdehyde (MDA), and proline in seedling increased with the decrease of the osmotic potential of PEG 6000 solution. The activities of peroxidase (POD), catalase (CAT), ascorbate peroxidase (APX), and glutathione reductase (GR) in seedling increased and then decreased with the decrease of osmotic potential of PEG 6000 solution. -0.1 MPa was the optimal osmotic potential of PEG 6000 solution simulated drought stress at germination stage for Job’s tears. The proline content and activities of POD and CAT were important mechanisms for the maintenance of drought resistance in Job’s tears seedling.

Isolating the Mutator Transposable Element Insertional Mutant Gene mio16 of Maize Using Double SelectedAmplification of Insertion Flanking Fragments (DSAIFF)

Mutator transposable element （Mu） has been used as an effective tool to clone maize （Zea mays L.） genes. One opaque endosperm mutant （miol6） was identified in a pool of Mu inserted mutants. A modified method, termed the double selected amplification of insertion flanking fragments （DSAIFF）, was employed to isolate the Mu flanking fragments （MFFs） of miol6. The target site duplications （TSDs） isolated from the Msp I and Mse I digested MFFs had a same 9-bp sequence and were confirmed to be the flanking sequence of one identically inserted gene. Co-segregation analysis suggested that the MFFs were associated with the mutant opaque endosperm, and miol6 was mapped in silico onto the physical position ranged from 229 965 021 to 229 965 409 bp of the maize chromosome 4.09 bin. The full-length cDNA of the wild-type gene was obtained by an RT-PCR primer-scanning technique, and Mio16 was found to putatively encode a homolog of the Arabidopsis MAP3K delta-1 protein kinase. RT-PCR result the mRNA expression of miol6 region anchored by primers Mu20 and af276 was not interrupted by Mu insertion. Further researches will be done to elucidate how the expression of miol6 is alternated by Mu insertion.

Evaluation on Inducible Effect of pOp6 Promoter in Transgenic Rice

In order to analyze the Os1bglu4 phenotype,the inducible promoter of the transgenic rice which knock-down the Os1bglu4 expression was assessed.The result showed that 30μM dexamethasone(DEX)had the stronger induction effect than10μM DEX byβ-Glucuronidase(GUS)staining.qRT-PCR further verified the Os1bglu4 gene deletion.The effect of DEX and its solvent absolute ethanol on seed development was measured,and no significant effect was observed.The conclusion is that final concentration of DEX at 30μM is suitable for pOp6 promoter induction.

Optimization of Quantitative Real-time PCR System on Amplification of Beta-glucosidase Gene Os1bglu4

[Objective] This study aimed to establish a quantitative real-time PCR(qRT-PCR) system for detecting the expression of rice beta-glucosidase gene Os1bglu4. [Method] The PCR was conducted with SYBR Green I method, using the primers of reference gene actin or ubiquitin. [Result] Actin was more suitable to be the reference gene than ubiquitin. More accurate results were obtained when the 100 ng cDNA template was added at a large volume and a lower concentration.The primer concentration in the range from 0.2 to 0.8 μmol/L we set had no significant influence on the results, so, 0.4 μmol/L was selected as the optimal primer concentration in this study. The amplification efficiency was greatly reduced when the annealing temperature was set at 64 ℃, therefore, annealing temperature was set at 60 ℃. Compared with the reaction system of 25 μl, the fluorescence intensity was significantly lower but the CT value did not change greatly in 10 μl system.So, the 10 μl reaction system was selected, which significantly reduces the research costs for the detection of a large amount of samples in future study.