Abstract
Rice is the model plant system for monocots and the sequencing of its genome has led to the identification of a vast array of genes for characterization. The tedious and time-consuming effort of raising rice transgenics has significantly delayed the pace of rice research. The lack of highly efficient transient assay protocol for rice has only added to the woes which could have otherwise helped in rapid deciphering of the functions of genes. Here, we describe a technique for efficient transient gene expression in rice seedlings. It makes use of co-cultivation of 6-day-old rice seedlings with Agrobacterium in the presence of a medium containing Silwet® L-77, acetosyringone and glucose. Seedlings can be visualized 9 days after co-cultivation for transient expression. The use of young seedlings helps in significantly reducing the duration of the experiment and facilitates the visualization of rice cells under the microscope as young leaves are thinner than mature rice leaves. Further, growth of seedlings at low temperature, and the use of surfactant along with wounding and vacuum infiltration steps significantly increases the efficiency of this protocol and helps in bypassing the natural barriers in rice leaves, which hinders Agrobacterium-based transformation in this plant. This technique, therefore, provides a shorter, efficient and cost-effective way to study transient gene function in intact rice seedling without the need for a specialized device like particle gun.
Highlights
Rice is an important cereal crop plant, which is the staple food of Asia and part of the pacific
To have a time-efficient protocol, the rice seedling part to be used for transient experiments should emerge within 1 week and should stay alive for a reasonable number of days for the transformed gene to express in sufficient quantity
The presence of thick waxy cuticle and high silica content has been a major barrier in standardizing methods of transient expression in rice plants
Summary
Rice is an important cereal crop plant, which is the staple food of Asia and part of the pacific. According to FAO, there is a thin line of rice self-sufficiency in many countries and the increasing population pressure is making this line disappear fast (Sainsbury and Lomonossoff, 2014). The ultimate aim of rice research is to increase production using less land, less manpower, less water and fewer pesticides. To achieve this aim, a basic understanding of this plant is essential, which requires in depth knowledge of the functions of different genes and their interactions. One way to study the function of rice genes is to express them in plants through stable or transient expression (Sainsbury and Lomonossoff, 2014). A transient expression system is considered ideal if it is technologically simple, cost-effective, robust, high
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