Agroinfiltration of intact leaves as a method for the transient and stable transformation of saponin producing Maesa lanceolata
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A method has been developed to genetically transform the medicinal plant Maesa lanceolata. Initially, we tested conditions for transient expression of GFP-bearing constructs in agroinfiltrated leaves. Leaf tissues of M. lanceolata were infiltrated with Agrobacterium tumefaciens carrying a nuclear-targeted GFP construct to allow the quantification of the transformation efficiency. The number of transfected cells was depended on the bacterial density, bacterial strains, the co-cultivation time, and presence of acetosyringone. The transient transformation assay generated the highest ratio of transfected cells over non-transfected cells upon 5 days post-infiltration using A. tumefaciens strain LBA4404 at an OD600 = 1.0 in the presence of 100 μM acetosyringone and in the absence of a viral suppressor construct. In a second series of experiments we set up a stable transformation protocol that resulted in the regeneration of kanamycin-resistant plants expressing nuclear GFP. This transformation protocol will be used to introduce overexpression and RNAi constructs into M. lanceolata plants that may interfere with triterpenoid saponin biosynthesis.
Key message We have developed a transformation protocol for saponin producing Maesa lanceolata. Using the protocol reported here, now we are able to generate the tools for the modification of saponin production.
KeywordsAgroinfiltration Transient gene expression Stable transformation Maesa lanceolata Saponin
The authors acknowledge David Baulcombe from Sainsbury Laboratory, Norwich, United Kingdom for providing the p19 construct. This research was funded by FWO-Flanders (project No. G.0014.08) and we thank the Directorate General of Higher Education, Ministry of Education and Culture, Republic of Indonesia for providing financial support for A.F.
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