5-Azacytidine promotes shoot regeneration during Agrobacterium-mediated soybean transformation.

Agrobacterium-mediated soybean transformation has been greatly improved in recent years, however the transformation efficiency is still low and highly genotype-dependent when compared to other species. Here, we characterized seventeen soybean genotypes based on their genetic transformation efficiencies, i.e., high and low, during Agrobacterium-mediated transformation. To reveal the molecular basis of this transformation difference, we constructed a highly efficient transient transgene expression system using soybean cotyledon protoplasts and then assess the methylation levels of promoter and coding regions of an EYFP (enhanced yellow fluorescent protein) gene introduced into the protoplast cultures of various soybean genotypes using BSP (bisulfite sequencing PCR). Increased methylation was found to be associated with the considerably decreased transfection efficiency (as percentage of EYFP fluorescent protoplasts) in low-efficacy genotypes as compared with those in high-efficacy on three DAT (day after transfection). 5-Azacytidine (5-Azac), a demethylating reagent commonly applied in epigenetic researches, significantly improved the transient transfection efficiency and transgene expression level in low-efficiency genotypes. Furthermore, the shoot regeneration efficiency in low-efficiency genotypes was substantially increased by 5-Azac treatment in an Agrobacterium-mediated soybean transformation system. Taken together, we concluded that lower methylation level in transgene contributed to enhanced shoot regeneration in Agrobacterium-mediated soybean transformation.

Comparison of Soybean Transformation Efficiency and Plant Factors Affecting Transformation during the Agrobacterium Infection Process.

The susceptibility of soybean genotype to Agrobacterium infection is a key factor for the high level of genetic transformation efficiency. The objective of this study is to evaluate the plant factors related to transformation in cotyledonary nodes during the Agrobacterium infection process. This study selected three genotypes (Williams 82, Shennong 9 and Bert) with high transformation efficiency, which presented better susceptibility to Agrobacterium infection, and three low transformation efficiency genotypes (General, Liaodou 16 and Kottman), which showed a relatively weak susceptibility. Gibberellin (GA) levels and soybean GA20ox2 and CYP707A2 transcripts of high-efficiency genotypes increased and were higher than those of low-efficiency genotypes; however, the opposite performance was shown in abscisic acid (ABA). Higher zeatin riboside (ZR) content and DNA quantity, and relatively higher expression of soybean IPT5, CYCD3 and CYCA3 were obtained in high-efficiency genotypes. High-efficiency genotypes had low methyl jasmonate (MeJA) content, polyphenol oxidase (PPO) and peroxidase (POD) activity, and relatively lower expression of soybean OPR3, PPO1 and PRX71. GA and ZR were positive plant factors for Agrobacterium-mediated soybean transformation by facilitating germination and growth, and increasing the number of cells in DNA synthesis cycle, respectively; MeJA, PPO, POD and ABA were negative plant factors by inducing defence reactions and repressing germination and growth, respectively.

Alleviation of exogenous oligochitosan on wheat seedlings growth under salt stress.

Hydroponic experiments were carried out to study the role of oligochitosan in enhancing wheat (Triticum aestivum L.) resistance to salt stress. Data were collected on plant biomass, chlorophyll content, photosynthetic rate (P (n)), stomatal conductance (g (s)), proline content, antioxidant enzyme activities, and malondialdehyde (MDA) content. Under 150 mM salt stress, plant growth was significantly inhibited. Shoot length, root length, and dry weight were sharply reduced by 26%, 31%, and 20%, respectively, of the control. Superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD) were increased by 28%, 13%, and 26%, respectively, of the control and MDA content largely accumulated, which was 1.5-fold of the control. However, 0.0625% oligochitosan pretreatment alleviated the adverse effects of salt stress, which was reflected by increasing root length, shoot length, dry weight, chlorophyll content, P (n,) and g (s). Furthermore, it also showed that oligochitosan pretreatment significantly increased antioxidant enzyme (SOD, CAT and POD) activities, and reduced MDA content in leaves. Meanwhile, the accumulation of proline was markedly accelerated. The results indicated that oligochitosan pretreatment ameliorated the adverse effects and partially protected the seedlings from salt stress during the following growth period.

[Specific promoters used in plant gene engineering].

The choice of specific promoters used within a transgene construct is a vital strategy to achieve the transgene regulation in the temporal, spatial and measurable manner. The strategy has been widely used in diverse aspects of plant gene engineering, such as quality improvement, resistance breeding and bioreactor. In this paper, we describe the structure feature, classification and research method of the specific promoter and its application progresses in plant gene engineering.