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Asian Journal of Biotechnology
  Year: 2014 | Volume: 6 | Issue: 1 | Page No.: 1-14
DOI: 10.3923/ajbkr.2014.1.14
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Optimizations of Laboratory-scale Vacuum-assisted Agroinfiltration for Delivery of a Transgene in Nicotiana benthamiana

Hwei-San Loh and Samson Baranzan Wayah

Agrobacterium-mediated transient gene transfer or agroinfiltration is a method to induce transient expression of heterologous proteins in plants. It offers speed, low cost and convenience when compared to traditional plant transformation. This study was aimed to set up a simple but feasible laboratory-scale vacuum agroinfiltration system. Basically, the procedures involved whole plant immersion into transformed Agrobacterium tumefaciens suspension and pressure application via vacuum drawing the agrobacteria into the plant tissues homogeneously. Optimizations of its parameters such as vacuum pressure and treatment duration towards the expression activity of a reporter, Green Fluorescence Protein (GFP) were conducted. The transformed A. tumefaciens strain LBA4404 carrying the recombinant plant viral vector, pEAQ-HT-GFP was infiltrated into the leaves of N. benthamiana via vacuum mode. In general, vacuum pressure and duration of infiltration had significant effects (p<0.05) on transient expression of GFP. Both fluorescence microscopy and Western blotting showed that vacuum agroinfiltration at 600 mmHg for 3 min had the highest and 400 mmHg for 2 min had the lowest mean GFP expression levels. This modified vacuum agroinfiltration approach utilized only distilled water and A. tumefaciens as components of the infiltration solution had substantially reduced the chemical cost. In conclusion, a simple and cost-effective laboratory-scale vacuum-assisted agroinfiltration system has been developed for recombinant protein production in planta.
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How to cite this article:

Hwei-San Loh and Samson Baranzan Wayah, 2014. Optimizations of Laboratory-scale Vacuum-assisted Agroinfiltration for Delivery of a Transgene in Nicotiana benthamiana. Asian Journal of Biotechnology, 6: 1-14.

DOI: 10.3923/ajbkr.2014.1.14






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