4 Notes
1. The growth vigor of the plants is one of the key factors determining a high success rate in grafting. To grow Arabidopsis
rootstocks, we used round pots 5.5 cm in diameter and 5 cm
high, which can be placed under a dissecting microscope. Up to
3 seeds are directly sowed or transplanted after germination. To
grow the scions, 9 seedlings are transplanted to a 9 Â 9-cm
square pot.
2. The flowering time of LD- or SD-grown Arabidopsis occurs
differently. Because LD-grown Arabidopsis flowers much earlier, the optimized stage for epicotyl grafting is limited. Under
our LD conditions, the grafted plants usually flowers 2 weeks
after grafting, which provides sufficient time to examine mobile
signals. Conversely, SD-grown Arabidopsis exhibit longer vegetative growth; thus, the grafting can be conducted at a later
stage to ensure robustness of the rootstocks.
3. Keeping part of the leaves on the scions increases the size of the
scions, which improves the success of grafting. In addition, the
cut on the leaves provides a useful marker to distinguish scions
from rootstocks after prolonged growth of grafted plants,
especially when no phenotypic markers can be used to distinguish scions from rootstocks.
4. It is important to ensure that the cutting surface on the hypocotyl of the scions is flat and intact. Always use the sharp edge of
the razor blade to make a clear cut and avoid squashing the
hypocotyl (i.e., gently slice the blade through the hypocotyl of
the scion without pressing it with the blade). To ensure that the
blade is sharp, use a fresh razor blade.
5. The base of the petiole and the hypocotyl is not vertically
aligned. To ensure that the insect pin is inserted in the middle
of the hypocotyl, exploit the flexibility of the insect pins to
adjust the angle of the insertion. Do not move insect pins
back and forth in the hypocotyl, which may severely damage
the plants.
6. To assemble the scions with rootstocks, insert the
scion-containing pin vertically onto the cutting surface of the
rootstocks and avoid protruding the insect pin out of the rootstocks, which may cause further damage and reduce the success
rate of grafting. We usually let the insect pin jut out of the
hypocotyl of scions about 1 mm and slowly insert the pin into
rootstocks. Once the scions and rootstocks are tightly attached,
you will feel resistance from the pin. Avoid moving the pin back
and forth to avoid damaging the rootstocks.
Arabidopsis Non-sterile Grafting
117
1. The growth vigor of the plants is one of the key factors determining a high success rate in grafting. To grow Arabidopsis
rootstocks, we used round pots 5.5 cm in diameter and 5 cm
high, which can be placed under a dissecting microscope. Up to
3 seeds are directly sowed or transplanted after germination. To
grow the scions, 9 seedlings are transplanted to a 9 Â 9-cm
square pot.
2. The flowering time of LD- or SD-grown Arabidopsis occurs
differently. Because LD-grown Arabidopsis flowers much earlier, the optimized stage for epicotyl grafting is limited. Under
our LD conditions, the grafted plants usually flowers 2 weeks
after grafting, which provides sufficient time to examine mobile
signals. Conversely, SD-grown Arabidopsis exhibit longer vegetative growth; thus, the grafting can be conducted at a later
stage to ensure robustness of the rootstocks.
3. Keeping part of the leaves on the scions increases the size of the
scions, which improves the success of grafting. In addition, the
cut on the leaves provides a useful marker to distinguish scions
from rootstocks after prolonged growth of grafted plants,
especially when no phenotypic markers can be used to distinguish scions from rootstocks.
4. It is important to ensure that the cutting surface on the hypocotyl of the scions is flat and intact. Always use the sharp edge of
the razor blade to make a clear cut and avoid squashing the
hypocotyl (i.e., gently slice the blade through the hypocotyl of
the scion without pressing it with the blade). To ensure that the
blade is sharp, use a fresh razor blade.
5. The base of the petiole and the hypocotyl is not vertically
aligned. To ensure that the insect pin is inserted in the middle
of the hypocotyl, exploit the flexibility of the insect pins to
adjust the angle of the insertion. Do not move insect pins
back and forth in the hypocotyl, which may severely damage
the plants.
6. To assemble the scions with rootstocks, insert the
scion-containing pin vertically onto the cutting surface of the
rootstocks and avoid protruding the insect pin out of the rootstocks, which may cause further damage and reduce the success
rate of grafting. We usually let the insect pin jut out of the
hypocotyl of scions about 1 mm and slowly insert the pin into
rootstocks. Once the scions and rootstocks are tightly attached,
you will feel resistance from the pin. Avoid moving the pin back
and forth to avoid damaging the rootstocks.
Arabidopsis Non-sterile Grafting
117