biologia plantarum

International journal on Plant Life established by Bohumil Němec in 1959

Biologia plantarum 48:367-374, 2004 | DOI: 10.1023/B:BIOP.0000041088.62614.76

Production of Transgenic Soybean Plants with Two Anti-Fungal Protein Genes Via Agrobacterium and Particle Bombardment

H.Y. Li1, Y.M. Zhu2, Q. Chen1, R.L. Conner3, X.D. Ding2, J. Li2, B.B. Zhang2
1 Agriculture and Agri-Food Canada, Lethbridge Research Centre, Lethbridge, AB, Canada, e-mail
2 College of Life Science, Northeast Agricultural University, Harbin, China
3 Agriculture and Agri-Food Canada, Morden Research Station, Morden MB, Canada

Utilizing either Agrobacterium-mediated transformation or particle bombardment we obtained transgenic soybean [Glycine max (L.) Merr.] plants expressing the chitinase gene (chi) and the barley ribosome-inactivating protein gene (rip). Six regenerated plants were grown to maturity and set seed. The identification of transgenic soybean plants that co-integrated the two anti-fungal protein genes was determined by polymerase chain reaction (PCR) and Southern blot analysis. Protein detection from the soybean leaves demonstrated the expression of the chitinase (CHI) and the ribosome-inactivating protein (RIP) in the six R0 transformants. Soybean cotyledonary nodes were transformed using the bivalent plant expression vector pBRC containing chi and rip both driven by the CaMV 35S double promoter. Following vacuum (0.06 MPa) infiltration treatment of the tissue for 5 min, Agrobacterium was co-cultivated with the cotyledonary nodes for 3 d on MSB medium (MS salts and B5 vitamins) (pH 5.2), the transformation frequency reached a maximum of 1.33 %. The chi and rip genes were present in all the transgenic plants. Co-bombardment of immature cotyledons with plasmids pBchE (encoding chi) and pARIP (encoding rip) resulted in a maximum transformation frequency of 0.52 % with a 50 % co-integration rate. Our results demonstrate efficient co-transformation of multiple genes in soybean.

Keywords: barley ribosome-inactivating protein gene; bean chitinase gene; CHI and RIP expression; PCR; Southern blot
Subjects: Agrobacterium-mediated transformation; Agrobacterium tumefaciens; barley, ribosome-inactivating protein gene; chitinase; cytokinins; Glycine max; nutrient medium, B5 (Gamborg); nutrient medium, Murashige and Skoog; nutrient medium, YEP; polymerase chain reaction (PCR); Southern blot analysis; soybean, transgenic; transformation, Agrobacterium-mediated; transformation, particle bombardment; transgenic plants

Published: September 1, 2004  Show citation

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Li, H.Y., Zhu, Y.M., Chen, Q., Conner, R.L., Ding, X.D., Li, J., & Zhang, B.B. (2004). Production of Transgenic Soybean Plants with Two Anti-Fungal Protein Genes Via Agrobacterium and Particle Bombardment. Biologia plantarum, 48(3), 367-374. doi: 10.1023/B:BIOP.0000041088.62614.76
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