A pin-fasten grafting method provides a non-sterile and highly efficient method for grafting Arabidopsis at diverse developmental stages

Background: Higher plants have evolved sophisticated communication systems to integrate environmental stimuli into their developmental programs. Grafting provides a powerful technique to examine transportation and systemic effects of mobile molecules. In Arabidopsis, many grafting approaches have been developed to investigate systemic molecules. However, these methods are usually limited to specific developmental stages or require sterilized conditions. To broaden the application of grafting for examining systemic signals at diverse developmental stages, we developed an Arabidopsis pin-fasten grafting method with insect pins used to assemble stocks and scions. Results: We report the step-by-step protocol of Arabidopsis pin-fasten grafting. Arabidopsis wild-type or gl1-1 plants were grown under long-or short-day conditions. Insect pins were inserted into gl1-1 scions at different developmental stages for grafting onto epicotyls or hypocotyls of stocks. Successfully grafted scions with newly developed glabrous leaves were observed at 14 days after grafting. Further longitudinal sections of the graft union showed well-connected vascular tissues between grafted plants. Use of fluorescent phloem-limited dye carboxyfluorescein diacetate in grafted plants demonstrated a symplastic connection established at 6 days after grafting and almost fully developed at 8 days. Conclusions: Our method provides a simple and robust approach to grafting Arabidopsis at different developmental stages. Sterilized conditions are not required, which greatly improves the success of grafting and plant growth.