Assessment of factors influencing the Agrobacterium-mediated in planta seed transformation of brinjal (Solanum melongena L.).

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).

• Authors: Kondeti Subramanyam, Manoharan Rajesh, Balusamy Jaganath, Amirthalingam Vasuki, Jeevaraj Theboral, Dhandapani Elayaraja, Sivabalan Karthik, Markandan Manickavasagam, Andy Ganapathi
• Journal: Applied biochemistry and biotechnology (Appl Biochem Biotechnol) Vol. 171 Issue 2 Pg. 450-68 (Sep 2013) ISSN: 1559-0291 [Electronic] United States
• PMID: 23852797 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Acetophenones
• Surface-Active Agents
• acetosyringone
• Glucuronidase

Topics

• Acetophenones (pharmacology)
• Agrobacterium (genetics)
• Coculture Techniques
• Genetic Engineering (methods)
• Genotype
• Germination
• Glucuronidase (genetics, metabolism)
• Seeds (genetics, growth & development)
• Solanum melongena (drug effects, enzymology, genetics, growth & development)
• Sonication
• Surface-Active Agents (pharmacology)
• Time Factors
• Transformation, Genetic (drug effects)
• Transgenes (genetics)
• Vacuum