Background

Grafting has served as a method to assemble two different types of plants, not only in agriculture but also in the field of plant science. Although plants do not have a nervous system, many studies using grafting experiments have demonstrated that plants properly integrate information from the entire body (Wang, 2011; Goldschmidt, 2014; Notaguchi and Okamoto, 2015; Thomas and Frank, 2019). In response to changes in their surrounding environments, plants generate information signals locally at the site of reception and transport signaling molecules systemically to transmit information. Inside plants, organ-to-organ communication is achieved via vascular tissues using a wide variety of molecules such as phytohormones, small RNAs, mRNAs, and proteins.

To study systemic signaling, especially between shoots and roots, micrografting, also known as seedling grafting, has been used. Micrografting is a method for assembling a scion and a rootstock that are cut out of seedlings from different plants. Since the first micrografting technique in the model plant Arabidopsis was demonstrated by Turnbull et al. (2002), micrografting has been widely used in a great number of studies and has contributed to the identification of molecules involved in systemic signaling such as flowering, defense against biotic infection, nutrient absorption and allocation, and drought stress response (Tsutsui and Notaguchi, 2017).

However, the micrografting technique requires dexterous operation and practice to achieve a high rate of success. Several papers have developed improved methods using modified procedures and equipment (Notaguchi et al., 2009), an agar-base support (Marsch-Martínez et al., 2013), and a fine pin to align the scion and the stock (Huang and Yu, 2015); nevertheless, micrografting remains challenging for beginners. To overcome the difficulties in conventional micrografting methods and make this technique accessible to those who study systemic signaling, we recently developed a microdevice made of silicone rubber, called a micrografting chip (Tsutsui et al., 2020). The device was fabricated by Micro-Electro Mechanical Systems using poly(dimethylsiloxane) (PDMS). The micrografting chip supports the manipulation of entire micrografting processes and facilitates the generation of uniformly grafted plants. This method has been applied in studies on plant biology using Arabidopsis mutants (Tsutsui et al., 2020; Shinozaki et al., 2020; Kurotani et al., 2020) and molecular mechanisms of grafting (Notaguchi et al., 2020). Here, we show the procedure for micrografting using a micrografting chip, which is slightly modified in its structure from Tsutsui et al. (2020) (Figure 1).

Figure 1. Structure of a micrografting chip. A. Image of one unit of the micrografting chip. B. Layout of the unit of the micrografting chip. C. Images of the micrografting chip (top) and its cover (bottom). D. Layouts of the micrografting chip (top) and its cover (bottom).