3. Use Iris scissors to cut a scion from the base of the hypocotyl.
Use scissors to trim the mature leaves from 1/3 of the leaf
blade (Fig. 1E; see Note 3). Place the leaf-removed scion under
the second dissecting microscope. Cut the hypocotyl of the
scion from 0.1 cm below the cotyledons to create a flat surface
on a hypocotyl by using a double-edged stainless steel razor
blade (see Note 4).
4. Use a high-precision tweezer to pick a 0.1-mm insect pin and
another tweezer to hold the scion. To avoid damaging the
scions, gently buckle up the scions from the axil of the leaves
with a tweezer (Fig. 1f). Under a dissecting microscope, insert
the insect pin from the base of the petiole through the hypocotyl and let the pin protrude out the hypocotyl about 1 mm
(Fig. 1g; see Note 5).
5. Assemble the scions with rootstocks by vertically inserting a
scion-containing pin into the flat surface of an apex-removed
rootstock (Fig. 1h; see Note 6).
6. Transfer the grafted plants to a tray, cover the tray with a lid to
retain humidity for 1 week. One week after grafting, gradually
open the lid and transfer the grafted plants to a growth chamber with normal growth conditions (see Note 7).
3.2 Pin-Fasten
Hypocotyl Grafting
1. Arabidopsis seeds are grown on soil under SD conditions for
2 months (see Note 8). To prepare rootstocks, use Iris scissors
to remove 2–3 leaves near the yet-to-be-grafted area on the
hypocotyl of SD-grown Arabidopsis to expose the hypocotyl
and to prevent shading the scions (see Note 9).
2. Place the pot of rootstocks under a dissecting microscope, use a
sharp microsurgical knife to scrape the surface of the hypocotyl
of rootstocks to remove the epidermis. The diameter of the
epidermis-removed area should be equal to the diameter of the
scion hypocotyl.
3. To prepare scions, use Iris scissors to cut the plants from the
base of the hypocotyl, remove mature leaves but keep 3–5
young leaves on the scions (see Note 10). Cut the hypocotyl
of the scion from 0.1 cm below the cotyledons by using a
double-edged stainless steel razor blade. Insert a 0.1-mm insect
pin into the scion as described in Subheading 3.1, step 4.
4. Assemble the plants by firmly inserting the scions to the
epidermis-removed hypocotyl of rootstocks. Transfer the
grafted plants to a tray, cover with a lid to keep humidity for
1 week. One week after grafting, gradually open the lid and
transfer the grafted plants to a growth chamber with normal
growth conditions (see Note 11).
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Nien-Chen Huang and Tien-Shin Yu
Use scissors to trim the mature leaves from 1/3 of the leaf
blade (Fig. 1E; see Note 3). Place the leaf-removed scion under
the second dissecting microscope. Cut the hypocotyl of the
scion from 0.1 cm below the cotyledons to create a flat surface
on a hypocotyl by using a double-edged stainless steel razor
blade (see Note 4).
4. Use a high-precision tweezer to pick a 0.1-mm insect pin and
another tweezer to hold the scion. To avoid damaging the
scions, gently buckle up the scions from the axil of the leaves
with a tweezer (Fig. 1f). Under a dissecting microscope, insert
the insect pin from the base of the petiole through the hypocotyl and let the pin protrude out the hypocotyl about 1 mm
(Fig. 1g; see Note 5).
5. Assemble the scions with rootstocks by vertically inserting a
scion-containing pin into the flat surface of an apex-removed
rootstock (Fig. 1h; see Note 6).
6. Transfer the grafted plants to a tray, cover the tray with a lid to
retain humidity for 1 week. One week after grafting, gradually
open the lid and transfer the grafted plants to a growth chamber with normal growth conditions (see Note 7).
3.2 Pin-Fasten
Hypocotyl Grafting
1. Arabidopsis seeds are grown on soil under SD conditions for
2 months (see Note 8). To prepare rootstocks, use Iris scissors
to remove 2–3 leaves near the yet-to-be-grafted area on the
hypocotyl of SD-grown Arabidopsis to expose the hypocotyl
and to prevent shading the scions (see Note 9).
2. Place the pot of rootstocks under a dissecting microscope, use a
sharp microsurgical knife to scrape the surface of the hypocotyl
of rootstocks to remove the epidermis. The diameter of the
epidermis-removed area should be equal to the diameter of the
scion hypocotyl.
3. To prepare scions, use Iris scissors to cut the plants from the
base of the hypocotyl, remove mature leaves but keep 3–5
young leaves on the scions (see Note 10). Cut the hypocotyl
of the scion from 0.1 cm below the cotyledons by using a
double-edged stainless steel razor blade. Insert a 0.1-mm insect
pin into the scion as described in Subheading 3.1, step 4.
4. Assemble the plants by firmly inserting the scions to the
epidermis-removed hypocotyl of rootstocks. Transfer the
grafted plants to a tray, cover with a lid to keep humidity for
1 week. One week after grafting, gradually open the lid and
transfer the grafted plants to a growth chamber with normal
growth conditions (see Note 11).
116
Nien-Chen Huang and Tien-Shin Yu