Agrobacterium-mediated transformation of Brassica napus and Brassica oleracea
- 17 January 2008
- journal article
- Published by Springer Science and Business Media LLC in Nature Protocols
- Vol. 3 (2), 181-189
- https://doi.org/10.1038/nprot.2007.527
Abstract
Agrobacterium-mediated transformation is widely used for gene delivery in plants. However, commercial cultivars of crop plants are often recalcitrant to transformation because the protocols established for model varieties are not directly applicable to them. The genus Brassica includes the oil seed crop, canola (B. napus), and vegetable crop varieties of Brassica oleracea, including cauliflower, broccoli and cabbage. Here, we describe an efficient protocol for Agrobacterium-mediated transformation using seedling explants that is applicable to various Brassica varieties; this protocol has been used to genetically engineer commercial cultivars of canola and cauliflower in our laboratory. Young seedling explants are inoculated with Agrobacterium on the day of explant preparation. Explants are grown for 1 week in the absence of a selective agent before being transferred to a selective medium to recover transgenic shoots. Transgenic shoots are subjected to an additional round of selection on medium containing higher levels of the selective agent and a low-carbohydrate source; this helps to eliminate false-positive plants. Use of seedling explants offers flexible experiment planning and a convenient explant source. Using this protocol, transgenic plants can be obtained in 2.5 to 3.5 months.Keywords
This publication has 31 references indexed in Scilit:
- Pod shatter‐resistant Brassica fruit produced by ectopic expression of the FRUITFULL genePlant Biotechnology Journal, 2005
- Agrobacterium-mediated transformation and generation of male sterile lines of Australian canolaAustralian Journal of Agricultural Research, 2005
- Crop Improvement through Modification of the Plant's Own GenomePlant Physiology, 2004
- In vitro shoot regeneration from commercial cultivars of Australian canola (Brassica napus L.)Australian Journal of Agricultural Research, 2004
- Transgenic organisms—time for conceptual diversification?Nature Biotechnology, 2003
- Transient β-gus and gfp Gene Expression and Viability Analysis of Microprojectile Bombarded Microspores of Brassica napus L.Journal of Plant Physiology, 2000
- Comparison of shoot regeneration potential from seedling explants of Australian cauliflower (Brassica oleracea var. botrytis) varietiesAustralian Journal of Agricultural Research, 1998
- Genetic transformation of BrassicaPlant Breeding, 1996
- Electroporation of rapeseed protoplasts - transient and stable transformationPhysiologia Plantarum, 1993
- A Revised Medium for Rapid Growth and Bio Assays with Tobacco Tissue CulturesPhysiologia Plantarum, 1962