Agrobacterium -mediated transformation of promising oil-bearing marine algae Parachlorella kessleri
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Agrobacterium-mediated transformation of promising oil-bearing marine algae Parachlorella kessleri Jayant Pralhad Rathod • Gunjan Prakash Reena Pandit • Arvind M. Lali
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Received: 17 April 2013 / Accepted: 23 September 2013 / Published online: 6 October 2013 Ó Springer Science+Business Media Dordrecht 2013
Abstract Parachlorella kessleri is a unicellular alga which grows in fresh as well as marine water and is commercially important as biomass/lipid feedstock and in bioremediation. The present study describes the successful transformation of marine P. kessleri with the help of Agrobacterium tumefaciens. Transformed marine P. kessleri was able to tolerate more than 10 mg l-1 hygromycin concentration. Co-cultivation conditions were modulated to allow the simultaneous growth of both marine P. kessleri and A. tumefaciens. For co-cultivation, P. kessleri was shifted from Walne’s to tris acetate phosphate medium to reduce the antibiotic requirement during selection. In the present study, the transfer of T-DNA was successful without using acetosyringone. Biochemical and genetic analyses were performed for expression of transgenes by GUS assay and PCR in transformants. Establishment of this protocol would be useful in further genetic modification of oil-bearing Parachlorella species. Keywords Parachlorella kessleri Agrobacteriummediated transformation Co-cultivation GUS assay hpt gene virC
J. P. Rathod G. Prakash R. Pandit A. M. Lali (&) DBT-ICT Centre for Energy Biosciences, Institute of Chemical Technology [University Under Section-3 of UGC Act 1956], Maharashtra Govt.’s Elite Status and Centre of Excellence, Grade ‘A’ by MHRD, Matunga, Mumbai, India e-mail: [email protected] A. M. Lali Department of Chemical Engineering, Institute of Chemical Technology [University Under Section-3 of UGC Act 1956], Maharashtra Govt.’s Elite Status and Centre of Excellence, Grade ‘A’ by MHRD, Matunga, Mumbai, India
Introduction The potential of algae as third-generation biofuels is being explored as the most effective alternative to fossil fuels. Mass-scale algal culturing can allow synergistic coupling of fuel generation, carbon sequestration and wastewater treatment resulting in sustainable energy conversion and utilisation. Algae biomass can be used directly as feedstock for biofuels or their high inherent oil content can be converted to biodiesel specifically (Brennan and Owende 2010; Chisti 2007). However, various biological as well as technological challenges need to be overcome to realise the true potential of any large-scale algal cultivation system (Hannon et al. 2010). Genetic modification can be one of the promising approaches to overcome the biological limitations of the algal system. In recent years, considerable advancements have been made on algal genetic modification (Hallmann 2007; Kiliana et al. 2011). However, this has been restricted mostly to the model system Chlamydomonas reinhardtii and few other freshwater species (Kumar et al. 2004; Cha et al. 2012; Kathiresan and Sarada 2009
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