A Unified Agrobacterium-Mediated Transformation Protocol for Alfalfa (Medicago sativa L.) and Medicago truncatula
Simplification of transformation procedures greatly improves work efficiency. In this chapter, we introduce a unified Agrobacterium-mediated transformation protocol that is used for both alfalfa (Medicago sativa L., Regen SY4D) and Medicago truncatula (ecotype R108). Whole trifoliates or leaflets are used as explants. Sonication is applied to enhance Agrobacterium infection and cytokinins are added to the medium to facilitate shoot regeneration. More than 90% transformation efficiency is achieved for alfalfa, while the relatively lower efficiency of up to 60% is obtained for M. truncatula, which depends on different selectable markers in the gene constructs. Transgenic plants are produced within 4–8 months with average timeline of 6 months. Using this unified protocol, the same types of media are used for both species which results in significant saving in time and resources.
Key wordsAlfalfa Medicago sativa Medicago truncatula Agrobacterium tumefaciens Genetic transformation Regeneration Transgenic plant Somatic embryogenesis
We would like to thank our previous and current group members Tim Hernandez, Yewei Wang, Steven Tudor, Mohammad Fereidouni, Guangming Li, Jianfei Yun, and Lishan Yang for their knowledge and suggestions in improving the procedure. We are also grateful to all the researchers and their principal investigators for their trust and support using the service.
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