Transformation of Stylosanthes spp. using Agrobacterium tumefaciens
1CSIRO Division of Tropical Crops and Pastures, CSIRO-UQ Plant Pathology Unit, Department of Botany, University of Queensland, 4067, St. Lucia, Australia.
Plant Cell Reports
|November 20, 2013
Summary
Agrobacterium tumefaciens incited tumours on Stylosanthes species. Gene transfer was successful in S. humilis using binary vectors, yielding kanamycin-resistant tissue expressing the neomycin phosphotransferase (NPT II) gene.
Area of Science:
- Plant molecular biology
- Plant biotechnology
- Agrobacterium-mediated transformation
Background:
- Stylosanthes species are important forage legumes.
- Agrobacterium tumefaciens is a natural plant pathogen used for genetic modification.
- Efficient gene transfer methods are crucial for crop improvement.
Purpose of the Study:
- To investigate Agrobacterium tumefaciens-mediated gene transfer in Stylosanthes species.
- To evaluate the suitability of binary vectors for transformation in S. humilis.
- To confirm the presence and expression of the neomycin phosphotransferase (NPT II) gene in transformed cells.
Main Methods:
- Infection of Stylosanthes leaf sections with Agrobacterium tumefaciens.
- Selection of kanamycin-resistant tissues.
- Molecular analysis (hybridization) to detect NPT II gene presence in DNA and RNA.
- Enzymatic assay to measure NPT II activity.
Main Results:
- Tumours were successfully induced on leaf sections of multiple Stylosanthes species.
- Kanamycin-resistant tumour tissue was obtained in S. humilis using binary vectors pGA472 and BIN6.
- The neomycin phosphotransferase (NPT II) gene was detected and expressed in resistant tumour cells.
- Teratomatous shoots expressing the NPT II gene were produced but could not be rooted.
Conclusions:
- Agrobacterium tumefaciens can be used for genetic transformation of Stylosanthes species.
- Binary vectors are effective for gene transfer in S. humilis.
- The NPT II gene confers kanamycin resistance and is expressed in transformed Stylosanthes cells, though regeneration of rooted plants remains a challenge.
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