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Assessment of Factors Influencing the Agrobacterium-mediated in planta Seed Transformation of Brinjal (Solanum melongena L.)

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Abstract

An efficient and reproducible in planta transformation method was developed for brinjal using seed as an explant. The brinjal seeds were infected with Agrobacterium tumefaciens EHA 105 harbouring pCAMBIA 1301-bar plasmid, and the transformants were selected against BASTA®. Several parameters influencing the in planta seed transformation such as pre-culture duration, acetosyringone concentration, surfactants, duration of sonication, vacuum pressure and vacuum duration have been evaluated. The putatively transformed (T 0) brinjal plants were screened by GUS histochemical analysis. Among the different combinations and concentrations tested, when the 18-h pre-cultured brinjal seeds were sonicated for 20 min and vacuum infiltered for 3 min at 500 mm of Hg in Agrobacterium suspension containing 100 μM acetosyringone, 0.2 % Silwett L-77 favoured the Agrobacterium infection and showed maximum transformation efficiency. Among the five brinjal varieties evaluated, Arka Samhitha showed maximum transformation efficiency at 45.66 %. The transgene was successfully transmitted to progeny plants (T 1) which was evidenced by GUS histochemical analysis, polymerase chain reaction and Southern hybridisation. The in planta protocol developed in the present study would be beneficial to transfer the economically and nutritionally important genes into different varieties of brinjal, and the transgenic brinjal plants can be produced in less time (approximately 27 days).

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Abbreviations

MS:

Murashige and Skoogs medium

B5:

Gamborg medium

gus :

β-glucuronidase gene

hptII:

Hygromycin phosphotransferase

CaMV 35S:

Cauliflower mosaic virus 35S promoter

GA3 :

Gibberellic acid

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Acknowledgements

The authors are thankful to Bharathidasan University, Tiruchirappalli, Tamil Nadu, India for providing lab facilities to carry out the present work. The corresponding author is thankful to University Grants Commission (UGC), Govt. of. India, for providing fellowship under UGC-BSR scheme. K. Subramanyam is thankful to Council of Scientific and Industrial Research (CSIR), New Delhi, India for providing Senior Research Fellowship to carry out his Doctoral work. J. Theboral is thankful to University Grants Commission (UGC), New Delhi, India for providing Moulana Azad Fellowship to carry out her Doctoral work.

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The authors declare that they have no competing interests.

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  1. Department of Biotechnology and Genetic Engineering, School of Biotechnology, Bharathidasan University, Tiruchirappalli, 620024, Tamil Nadu, India

    Kondeti Subramanyam, Manoharan Rajesh, Balusamy Jaganath, Amirthalingam Vasuki, Jeevaraj Theboral, Dhandapani Elayaraja, Sivabalan Karthik, Markandan Manickavasagam & Andy Ganapathi

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  1. Kondeti Subramanyam
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Subramanyam, K., Rajesh, M., Jaganath, B. et al. Assessment of Factors Influencing the Agrobacterium-mediated in planta Seed Transformation of Brinjal (Solanum melongena L.). Appl Biochem Biotechnol 171, 450–468 (2013). https://doi.org/10.1007/s12010-013-0359-z

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