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ESTABLISHMENT OF AGROBACTERIUM-MEDIATED BT GENE TRANSFORMATION SYSTEM IN MAT RUSH (JUNCUS EFFUSUS L.)

LING XU¹, ULLAH NAJEEB¹, WEI QI SHEN², GHULAM JILANI³, MUHAMMAD RASHEED¹ AND WEI JUN ZHOU^1*

Abstract: Agrobacterium-mediated genetic transformation was explored for inducing insect resistance in mat rush (Juncus effusus L.). Firstly, the callus regeneration system was established, and secondly the Agrobacterium tumefaciens strain EHA105 with plasmid pKUB carrying cryIA(b), hph and gus genes, coding for hygromycin phosphotransferase and β-glucuronidase, respectively, was used for transformation. Results indicated that 150 μM acetosyringone in both of the induction and co-cultivation media increased the frequency of GUS⁺ calli, and OD₆₀₀ value 0.4 of Agrobacterium enhanced the rate of GUS⁺ calli, significantly. The infection duration of 45-60 min, co-cultivation duration of 4 days, and infection in the 12^th or 16^th day were the best combination for transformation. Agrobacterium-treated plants achieved a transformation frequency of 9.56 % to the highest. Transient GUS expression and hygromycin-resistance tests were practicable and reliable for selecting the positive transformants. This transformation system was found suitable for manipulating the desired traits through transgenic techniques, as the transgenic Juncus effusus L. plants were produced from seeds to embryogenic calli, then to plantlets. The Agrobacterium - mediated transformation of Bt gene into mat rush with some optimizations is highly successful and reproducible as experienced from the established protocol.
 

¹Institute of Crop Science, Zhejiang University, Hangzhou 310029, China

²The University of Nottingham Ningbo, Ningbo 315100, China

³Department of Soil Science, PMAS Arid Agriculture University, Rawalpindi 46300, Pakistan