experiments. Mutant plants deficient in specific signals can be used
as background-free living materials to graft to different rootstocks
for examining the transport of mobile signals in vivo. However,
Arabidopsis is a rosette-type plant; the short internode and diminutive stature of plants greatly impedes manipulation with grafting.
In the past few years, many efforts have been made to develop
different grafting methods in Arabidopsis. In addition, the cell
division and vascular reconnection patterns in the Arabidopsis
graft junction have been established [4, 5]. The Arabidopsis inflorescence, which contains long internodes and sufficient diameter,
provides ideal material for grafting operation. In this method, the
primary inflorescence of scions is cut to create a wedge style and
fused with the inflorescence of rootstocks. A fragment of polyethylene tubes is used to provide physical support for the scions [6–
9]. With high successful rate and easy handling, Arabidopsis inflorescence grafting has become a reliable method to examine mobile
molecules [6, 8, 10]. However, this method is only used at the
reproductive stage where many developmental events have already
occurred. In addition, because of the poor growth vigor of scion
inflorescence, observing phenotypic changes after grafting is difficult. These drawbacks significantly impede the application of inflorescence grafting.
The most widely used Arabidopsis grafting is micrografting
[11]. In this method, a seedling in cotyledon stage is used as a
scion to graft onto a hypocotyl of rootstock that is grown under
sterile conditions. Different procedures have been developed to
assemble scions with rootstocks in this method [11–13]. Although
micrografting has been used to identify many mobile signals,
including long-distance signals in shoot branching [11], florigen
[14], and mobile miRNA or siRNA [15, 16], the technique
requires sterilized conditions and well-trained experts. Thus,
micrografting in many laboratories remains a challenge.
Here we described a simple and robust method for Arabidopsis
seedling grafting under a non-sterile condition [17]. An insect pin
is used to secure scions to rootstocks. With this method, soil-grown
Arabidopsis at different developmental stages can be used for grafting onto epicotyls or hypocotyls of rootstocks with high success
rate. This grafting method has been successfully used to investigate
mobile signals in floral initiation [18, 19].
2 Materials
1. Long-day (LD) or short-day (SD) soil-grown Arabidopsis.
2. Two dissecting microscopes.
3. Microdissecting Vannas spring scissors, curved 80 mm, cutting
edge 2.5 mm.
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Nien-Chen Huang and Tien-Shin Yu
as background-free living materials to graft to different rootstocks
for examining the transport of mobile signals in vivo. However,
Arabidopsis is a rosette-type plant; the short internode and diminutive stature of plants greatly impedes manipulation with grafting.
In the past few years, many efforts have been made to develop
different grafting methods in Arabidopsis. In addition, the cell
division and vascular reconnection patterns in the Arabidopsis
graft junction have been established [4, 5]. The Arabidopsis inflorescence, which contains long internodes and sufficient diameter,
provides ideal material for grafting operation. In this method, the
primary inflorescence of scions is cut to create a wedge style and
fused with the inflorescence of rootstocks. A fragment of polyethylene tubes is used to provide physical support for the scions [6–
9]. With high successful rate and easy handling, Arabidopsis inflorescence grafting has become a reliable method to examine mobile
molecules [6, 8, 10]. However, this method is only used at the
reproductive stage where many developmental events have already
occurred. In addition, because of the poor growth vigor of scion
inflorescence, observing phenotypic changes after grafting is difficult. These drawbacks significantly impede the application of inflorescence grafting.
The most widely used Arabidopsis grafting is micrografting
[11]. In this method, a seedling in cotyledon stage is used as a
scion to graft onto a hypocotyl of rootstock that is grown under
sterile conditions. Different procedures have been developed to
assemble scions with rootstocks in this method [11–13]. Although
micrografting has been used to identify many mobile signals,
including long-distance signals in shoot branching [11], florigen
[14], and mobile miRNA or siRNA [15, 16], the technique
requires sterilized conditions and well-trained experts. Thus,
micrografting in many laboratories remains a challenge.
Here we described a simple and robust method for Arabidopsis
seedling grafting under a non-sterile condition [17]. An insect pin
is used to secure scions to rootstocks. With this method, soil-grown
Arabidopsis at different developmental stages can be used for grafting onto epicotyls or hypocotyls of rootstocks with high success
rate. This grafting method has been successfully used to investigate
mobile signals in floral initiation [18, 19].
2 Materials
1. Long-day (LD) or short-day (SD) soil-grown Arabidopsis.
2. Two dissecting microscopes.
3. Microdissecting Vannas spring scissors, curved 80 mm, cutting
edge 2.5 mm.
114
Nien-Chen Huang and Tien-Shin Yu