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

Huang NC, Yu TS - Plant Methods (2015)

Bottom Line: Grafting provides a powerful technique to examine transportation and systemic effects of mobile molecules.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.

View Article: PubMed Central - PubMed

Affiliation: Institute of Plant and Microbial Biology, Academia Sinica, Taipei, 11529 Taiwan.

ABSTRACT

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.

No MeSH data available.


Related in: MedlinePlus

Differentiation of gl1-1 scions in successfully grafted P35S-GUS transformant stocks. a, c Arabidopsis pin-fasten grafted plants at 14 days after grafting under LD conditions. Arabidopsis gl1-1 scions were grafted onto P35S-GUS transformant stocks. Trichrome-less leaves were developed from gl1-1 scions (white asterisks) and trichrome-containing leaves were from P35S-GUS transformant stocks. Insect pins are indicated by arrowheads. b, d Histochemical staining of Arabidopsis gl1-1 scions grafted onto P35S-GUS transformant stocks at 14 days after grafting. GUS activity was detected in stocks but not scions.
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Fig4: Differentiation of gl1-1 scions in successfully grafted P35S-GUS transformant stocks. a, c Arabidopsis pin-fasten grafted plants at 14 days after grafting under LD conditions. Arabidopsis gl1-1 scions were grafted onto P35S-GUS transformant stocks. Trichrome-less leaves were developed from gl1-1 scions (white asterisks) and trichrome-containing leaves were from P35S-GUS transformant stocks. Insect pins are indicated by arrowheads. b, d Histochemical staining of Arabidopsis gl1-1 scions grafted onto P35S-GUS transformant stocks at 14 days after grafting. GUS activity was detected in stocks but not scions.

Mentions: We further investigated the differentiation of scions and stocks on the grafted junction by histochemical staining (Figure 4). At 14 days after grafting, which gl1-1 scions were grafted onto P35S-GUS stocks under LD conditions, the newly differentiated glabrous leaves were observed from gl1-1 scions (Figure 4a, c). Histochemical assay showed GUS activity exclusively detected in P35S-GUS stocks but not glabrous scions (Figure 4b, d), which is consistent with GUS acting cell-autonomously [28].Figure 4


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

Huang NC, Yu TS - Plant Methods (2015)

Differentiation of gl1-1 scions in successfully grafted P35S-GUS transformant stocks. a, c Arabidopsis pin-fasten grafted plants at 14 days after grafting under LD conditions. Arabidopsis gl1-1 scions were grafted onto P35S-GUS transformant stocks. Trichrome-less leaves were developed from gl1-1 scions (white asterisks) and trichrome-containing leaves were from P35S-GUS transformant stocks. Insect pins are indicated by arrowheads. b, d Histochemical staining of Arabidopsis gl1-1 scions grafted onto P35S-GUS transformant stocks at 14 days after grafting. GUS activity was detected in stocks but not scions.
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4495618&req=5

Fig4: Differentiation of gl1-1 scions in successfully grafted P35S-GUS transformant stocks. a, c Arabidopsis pin-fasten grafted plants at 14 days after grafting under LD conditions. Arabidopsis gl1-1 scions were grafted onto P35S-GUS transformant stocks. Trichrome-less leaves were developed from gl1-1 scions (white asterisks) and trichrome-containing leaves were from P35S-GUS transformant stocks. Insect pins are indicated by arrowheads. b, d Histochemical staining of Arabidopsis gl1-1 scions grafted onto P35S-GUS transformant stocks at 14 days after grafting. GUS activity was detected in stocks but not scions.
Mentions: We further investigated the differentiation of scions and stocks on the grafted junction by histochemical staining (Figure 4). At 14 days after grafting, which gl1-1 scions were grafted onto P35S-GUS stocks under LD conditions, the newly differentiated glabrous leaves were observed from gl1-1 scions (Figure 4a, c). Histochemical assay showed GUS activity exclusively detected in P35S-GUS stocks but not glabrous scions (Figure 4b, d), which is consistent with GUS acting cell-autonomously [28].Figure 4

Bottom Line: Grafting provides a powerful technique to examine transportation and systemic effects of mobile molecules.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.

View Article: PubMed Central - PubMed

Affiliation: Institute of Plant and Microbial Biology, Academia Sinica, Taipei, 11529 Taiwan.

ABSTRACT

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.

No MeSH data available.


Related in: MedlinePlus