Fluorescent Soybean Hairy Root Construction and Its Application in the Soybean—Nematode Interaction: An Investigation

Background: The yield of soybean is limited by the soybean cyst nematode (SCN, Heterodera glycines). Soybean transformation plays a key role in gene function research but the stable genetic transformation of soybean usually takes half a year. Methods: Here, we constructed a vector, pNI-GmUbi, in an Agrobacterium rhizogenes-mediated soybean hypocotyl transformation to induce fluorescent hairy roots (FHRs). Results: We describe the operation of FHR-SCN, a fast, efficient and visual operation pathosystem to study the gene functions in the soybean-SCN interaction. With this method, FHRs were detected after 25 days in 4 cultivars (Williams 82, Zhonghuang 13, Huipizhiheidou and Peking) and at least 66.67% of the composite plants could be used to inoculate SCNs. The demographics of the SCN could be started 12 days post-SCN inoculation. Further, GmHS1pro-1 was overexpressed in the FHRs and GmHS1pro-1 provided an additional resistance in Williams 82. In addition, we found that jasmonic acid and JA-Ile increased in the transgenic soybean, implying that the resistance was mainly caused by affecting the content of JA and JA-Ile. Conclusions: In this study, we established a pathosystem, FHR-SCN, to verify the functional genes in soybeans and the SCN interaction. We also verified that GmHS1pro-1 provides additional resistance in both FHRs and transgenic soybeans, and the resistance may be caused by an increase in JA and JA-Ile contents.

Screening of Vietnamese soybean genotypes for Agrobacterium-mediated transgenic transformation

Soybean [Glycine max (L) Merr.] is one of the most important crops used for human food and animal feed globally. Transgenic soybean covers more than 74% of the global soybean production area, which is an achievement of genetically modified programs. The Agrobacterium-mediated method is commonly used for soybean transformation, but the efficiency of this method is affected by various factors including genotypes. Screening of the soybean genotypes suitable for Agrobacterium-infection and plant regeneration is the most important step to establish an efficient genetic transformation system. In this study, we screened thirty Vietnamese soybean genotypes including seventeen cultivated soybean genotypes (CSG) and thirteen local soybean genotypes (LCG) for shoot regeneration ability and transient infection via Agrobacterium tumefaciens method. Two CSG cultivars, DT22 and VX93, had significantly high efficiencies for shoot regeneration and transient infection compared with the control genotypes Jack and William 82. The shoot regeneration of DT22 and VX93 was 92.32% with 5.75 shoots/explant and 93.35% with 5.92 shoots/explant, respectively, whereas the control genotypes Jack and William 82 had 91.35% with 4.6 shoots/explant and 82.64% with 5.7 shoots/explant. Similarly, the transient infection of DT22 and VX93 was 84% and 86%, respectively, which was comparable with that of Jack (86%) William (82%). The success of transgenic development was confirmed by the β-Glucuronidase staining, PCR, and Basta leaf painting. The results indicated that cultivars DT22 and VX93 could be used for stable Agrobacterium-media transformation. Keywords: Soybean, Agrobacterium, transformation, transient infection, transgenic.

Improvement of Soybean Agrobacterium-Mediated Transformation Efficiency by Adding Glutamine and Asparagine into the Culture Media

As a genetically modified crop, transgenic soybean occupies the largest global scale with its food, nutritional, industrial, and pharmaceutical uses.Efficient transformation is a key factor for the improvement of genetically modified soybean. At present, the Agrobacterium-mediated method is primarily used for soybean transformation, but the efficiency of this method is still relatively low (below 5%) compared with rice (above 90%). In this study, we examined the influence of l-glutamine and/or l-asparagine on Agrobacterium-mediated transformation in soybean and explored the probable role in the process of Agrobacterium-mediated transformation. The results showed that when the amino acids l-glutamine and l-asparagine were added separately or together to the culture medium, the shoot induction frequency, elongation rate, and transformation frequency were improved. The combined effects of l-glutamine and l-asparagine were better than those of l-glutamine and l-asparagine alone. The 50 mg/L l-glutamine and 50 mg/L l-asparagine together can enhance the transformation frequency of soybean by attenuating the expression level of GmPRs (GmPR1, GmPR4, GmPR5, and GmPR10) and suppression of the plant defense response. The transgene was successfully transmitted to the T1 generation. This study will be useful in genetic engineering of soybean.

Transformasi Genetik Kedelai dengan Gen Proteinase Inhibitor II Menggunakan Teknik Penembakan Partikel

<p class="p1">An experiment was conducted at the Molecular Biology and Genetic Engineering Laboratory of BB-Biogen, Bogor with an objective to obtain transgenic soybean plants containing the <em>proteinase inhibitor </em>II (<em>pin</em>II) gene. The experiment consisted of three steps, i.e., optimalization of the soybean transformation technique using the <em>gus </em>gene; transformation of soybean using the <em>pin</em>II gene, and molecular analysis of the transformed soybean plants. Two type of explants (young embryo and cotyledon) were bombarded with <em>p</em>RQ6 plasmid containing the <em>gus </em>gene with the following treatment: Helium gas pressure (1100 psi and 1300 psi), shoot distance (5 and 7 cm), and number of bombardment (1x and 2x). The result of <em>gus </em>assay indicated that the best bombardment was done on young cotyledon explants with 1100 psi Helium pressure, shoot distance 5 cm, and 1x bombardment. Transformation of the soybean explant using the <em>pin</em>II gene (inside the <em>p</em>TWa plasmid) was conducted using the best bombardment treatment from the first activity. Two plants from c.v. Wilis (WP<span class="s1">1</span>, WP<span class="s1">2</span>) and three plants from c.v. Tidar (TP<span class="s1">1</span>, TP<span class="s1">2</span>, TP<span class="s1">3</span>) were recovered from regeneration and selection of the transformed explants. Molecular analysis of the regenerated plants using the PCR technique showed that only WP<span class="s1">2 </span>contained the <em>pin</em>II gene. This plant was fertile and will be used for further evaluation.</p>