**1. Introduction**

Grafting has become an effective practice in the production of high-value solanaceous and cucurbitaceous vegetables to help overcome biotic and abiotic stresses and improve crop productivity [1– 3]. Although grafting also has been used as a tool in plant physiology studies of Arabidopsis (*Arabidopsis thaliana* L.) [4], for accelerating the breeding work of common beans (*Phaseolus vulgaris* L.) [5], and for combating Verticillium wilt of globe artichoke (*Cynara cardunculus* L. subsp. *Scolymus*) [6], grafting in other vegetable species beyond Solanaceae and Cucurbitaceae is generally not practiced commercially. Interestingly, some attempts have been made to explore the feasibility of grafting vegetable plants in *Brassicaceae*. Oda et al. [7] tested inter-varietal, inter-specific, and inter-generic grafting among cabbage (*Brassica oleracea* L. var. *capitata*), kale (*Brassica oleracea* var. *sabellica*), kohlrabi (*Brassica oleracea* var. *gongylodes*), Chinese cabbage (*Brassica rapa* L. subsp. *pekinensis*), turnip (*Brassica rapa* subsp. *rapa*), Japanese mustard (Takana) (*Brassica juncea* L. var. *integrifolia*), and Japanese radish (*Raphanus sativus* L. var. *longipinnatus*) and obtained successful grafts. Particularly, an adhesive and hardener system was developed for making grafts between Chinese cabbage (scion) and turnip (rootstock) [8]. Recently, Chen et al. [9] evaluated the survival rate of cabbage grafted onto Chinese kale (*B. oleracea* Alboglabra group) rootstocks and assessed the feasibility of using grafting to improve cabbage head quality.

The effort of cruciferous vegetable grafting has not only broadened the potential use of vegetable grafting as a management tool, but also presents the possibility of creating a novel vegetable product with added value. In the case of grafted Chinese cabbage/turnip plants, the above-ground portion of Chinese cabbage—a leafy vegetable, and the below-ground portion of turnip—a root vegetable, can be harvested from the same plant. This type of rootstock–scion combination holds promise for space saving in small-scale intensive cultivation systems. Moreover, the grafted Chinese cabbage/turnip vegetable may possess added economic value with minimal waste, since many consumers may prefer not to eat turnip leaves and could be drawn to the novelty of this new product. However, the Chinese cabbage and turnip grafting study of Oda and Nakajima [8] only reported a 50% graft survival rate and observed restricted development of the Chinese cabbage head.

In this proof of concept study, we grafted the pac choi (*B. rapa* var. *chinensis*) scion onto the daikon radish (*R. sativus* var. *longipinnatus*) rootstock to generate a vegetable plant that produced a pac choi leafy vegetable above-ground and an edible daikon radish root below-ground. Pac choi and daikon radish are among specialty vegetables increasingly grown for local markets in the U.S. Although edible, daikon radish leaves are often discarded at consumption. Recent genetic studies supported the feasibility of making successful inter-generic grafts between *B. rapa* and *R. sativus*. Yang et al. [10] sequenced the chloroplast noncoding region and found that *R. sativus* was closely related to *B. rapa*/*oleracea* and proposed that *Raphanus* was derived from hybridization between *B. rapa*/*oleracea* and *B. nigra,* the two evolutionary lineages in the genus Brassica. Furthermore, the reciprocal hybridization between *R. sativus* and *B. rapa* has been proven viable [11]. Vigorous growth was also observed for most of the successful primary hybrids between *B. rapa* and *R. sativus* [12]. On the other hand, according to Tonosaki et al. [13], when hybridized with *R. sativus*, only one particular breeding line of *B. rapa* ('Shogoin-kabu') successfully produced hybrid seeds, whereas most other lines failed due to embryo breakdown.

By grafting the pac choi scion onto the daikon radish rootstock, the objectives of this pilot experiment were to examine the feasibility of developing successful grafts for harvesting both pac choi leaves and daikon radish taproot from the same plant, and to compare the growth and development of grafted plants with self-grafted and non-grafted pac choi and daikon radish plants.

#### **2. Materials and Methods**

Two experiments were carried out in this study. The first experiment was a pilot study to test the feasibility of grafting pac choi onto daikon radish. The second experiment was intended to provide a better understanding of above-ground growth and below-ground development of this unique scion-rootstock system over an extended post-grafting period of plant establishment. In both experiments, 'Bora King' (BK), a daikon radish with purple taproots (Johnny's Selected Seeds, Winslow, ME, USA) was used as the rootstock, while 'Mei Qing Choi' (MQ) pac choi (Johnny's Selected Seeds) was used as the scion. They were selected based on our preliminary study in which these two cultivars were found to be compatible for grafting and have similar hypocotyl diameters.

#### *2.1. Setup of the Pilot Experiment*

Pac choi and daikon radish were seeded on 7 and 13 November 2016, respectively. The pac choi was seeded 6 d earlier than the daikon radish in order to match the stem diameter of the seedlings at grafting, as the daikon radish germinated and emerged much quicker than the pac choi based on a preliminary seeding test. All the seeds were sown in 72-cell Speedling trays (Speedling Inc., Ruskin, FL, USA) and filled with Fafard-2 potting mix (Sun Gro Horticulture, Agawam, MA, USA) containing a mixture of peat moss, perlite, vermiculite, and dolomite lime. Plants were grown in a greenhouse at the University of Florida campus (Gainesville, FL, USA). Water-soluble fertilizer 20N-8.7P-16.7K (Jack's Classic; Jr Peters Inc., Allentown, PA, USA) was applied on 17 and 28 November at a nitrogen (N) concentration of 200 mg L<sup>−</sup>1.

Plants were grafted on 30 November 2016 (0 d after grafting (DAG)) using the splice grafting method [1]. Twenty-four plants were grafted using seedlings with the most consistent growth. With the purpose of ensuring consistent grafting quality, only a small number of plants were grafted in this pilot experiment after earlier attempts at practicing the grafting technique. The daikon radish seedlings were severed using a double edge razor blade at approximately a 45-degree angle below cotyledons to remove the shoots, with pac choi scions cut at the hypocotyl with the same angle just above the soil surface. The cut surfaces of the pac choi scion and the daikon radish seedlings with shoot removal were conjoined using a 1.5 mm silicone grafting clip (Johnny's Selected Seeds). Grafted plants were then placed in a healing chamber constructed by wrapping a metal shelving unit with thin plastic film (Uline Econo-Wrapper (0.02 mm), Uline corporation, Pleasant Prairie, WI, USA) in a temperature-controlled room with air temperature set at 23 ◦C and relative humidity (RH) at 99%. Light was provided by two, 54-watt T5 fluorescent lights (Philips Lighting Company, Somerset, NJ, USA) at a photosynthetic photon flux density (PPFD) of 56 μmol m−<sup>2</sup> s−<sup>1</sup> at seedling canopy level for 12 h each day. An additional plastic tray with a wet sheet of germination paper was placed inside the healing chamber to help maintain humidity.

From 5 DAG, the healing chamber was gradually cut open and the ambient RH setting was reduced to 60%. At 8 DAG, the plastic film was completely removed, and grafts remained exposed in the temperature-controlled room until 13 DAG. Water was applied to plants by filling the bottom of the tray for absorption. All the grafted plants were transferred to a greenhouse at 13 DAG, and graft survival rate was determined by counting the number of live and dead plants; only plants with turgid leaves were counted as living. At harvest, the number of surviving grafted plants was counted again for calculation of the final graft survival rate, as some plants severely declined following transplanting into pots.

At 16 DAG, surviving grafted plants were transplanted into 11.36 L black plastic pots (1200C; Hummert International, Earth City, MO, USA) filled with Fafard-2 soilless mix (Sun Gro Horticulture) for continued monitoring of the survival of the grafted plants. In addition, five plants of non-grafted 'Mei Qing Choi' and 'Bora King' were potted as controls. All the plants were placed on the greenhouse bench following a completely randomized design. Organic fertilizer MicroSTART60 3N-0.9P-2.5K (Perdue AgriRecycle, LLC., Seaford, DE, USA) was applied to each pot at the rate of 80 g/pot. Drip irrigation was used by placing one 1.89 L h−<sup>1</sup> emitter (Woodpecker pressure compensating junior dripper; Netafim USA, Fresno, CA, USA) in each pot; plants were watered once a day for 3 min. Irrigation increased to twice per day for 2 min each time starting at 56 DAG. Insecticidal soap (Safer Brand; Woodstream Corporation, Lancaster, PA, USA) was sprayed at 56 DAG and lacewing larvae (*Chrysoperla rufilabris* (Neuroptera: Chrysopidae); Rincon-Vitova Insectaries, Ventura, CA, USA) were released at 64 DAG for aphid control. The average day and night temperatures of the greenhouse during the plant growth were 22.8 ◦C and 16.5 ◦C, respectively.
