Research Note

Plant Cell, Tissue and Organ Culture

, Volume 94, Issue 3, pp 253-259

Sonication assisted Agrobacterium-mediated transformation enhances the transformation efficiency in flax (Linum usitatissimum L.)

  • Martina BeranováAffiliated withFaculty of Health and Social Studies, University of South Bohemia
  • , Slavomír RakouskýAffiliated withFaculty of Health and Social Studies, University of South BohemiaFaculty of Science, Department of Genetics, University of South Bohemia Email author 
  • , Zuzana VávrováAffiliated withFaculty of Health and Social Studies, University of South BohemiaFaculty of Science, Department of Genetics, University of South Bohemia
  • , Tomáš SkalickýAffiliated withFaculty of Health and Social Studies, University of South BohemiaFaculty of Science, Department of Genetics, University of South Bohemia

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Abstract

A sonication-assisted, Agrobacterium-mediated, co-cultivation technique was used in an attempt to increase the transformation efficiency of flax. Hypocotyls and cotyledons excised from about 10-day-old flax seedlings grown in vitro were placed into a 10 mM MgSO4 solution, and inoculated with an A. tumefaciens vector bearing the mgfp5-ER gene driven by the CaMV 35S promoter. The explants were subjected to pulses of ultrasound delivered by a sonicator apparatus (35 kHz) for 0–150 s and co-cultivated for 2 h at 27°C. The dried hypocotyls and cotyledons were grown on a selective MS medium to promote shoot regeneration. An electron microscopic study showed that the sonication treatment resulted in thousands of microwounds on and below the surface of the explants. A stereo microscope Leica MZ 12 equipped with a GFP adaptor was used to assess the infection and transformation of plant tissues in real time. After only 48 h and for at least 30 days after bacteria elimination, signs of transgene expression could be seen as a bright fluorescence. Our results show that treatment with ultrasound facilitates an enhanced uptake of plasmid DNA into the cells of flax hypocotyls and cotyledons and that its efficiency depends on the duration of the treatment and the frequency used. SAAT could be a promising tool for enhancing transformation efficiency in flax.

Keywords

CaMV 35S promoter Expression pattern Fluorescence Green fluorescent protein marker SAAT