U.S. flagAn official website of the United States government
                    Here's how you know **Official websites use .gov**
                            
                            A **.gov** website belongs to an official government organization in the United States.
 **Secure .gov websites use HTTPS**
                            
                            A **lock** (                            
                              A locked padlock                                                      
                            
                            ) or **https://** means you’ve safely connected to the .gov website. Share sensitive information only on official, secure websites.


Menu Agricultural Research Service
						
                    U.S. DEPARTMENT OF AGRICULTURE
 U.S. Dairy Forage Research Center: Madison, WI
 close



    Research

                    Research Projects
                                    Publications
                                    Collaborations
                                    News
                                    Software
                People

                    Staff Listing
                                    Careers


        


                    Archives
                        Presentations
                                    World Dairy Expo
                                    Newsletters and Research Summaries
                    Resources
                        General Resources
                                    Books
                                    Teacher's Resources
                                    New Research Dairy Updates
                                    Integrated Research at USDFRCSearch small
                    Search
            
            ARS Home
                                About ARS
                                Contact Us




ARS Home Midwest Area Madison, Wisconsin U.S. Dairy Forage Research Center Research Publications at this Location





- ARS National Programs
- Search for a scientific manuscript
- Search for a research project
- ARS Software
- Programs and Projects
- Collaborations
- Subjects of Investigation
- Publications
- IACUC Summaries - Milwaukee, WI Aquaculture
- IACUC Summaries - US Dairy Forage Research Center

Title:   Author  item
                                            Sullivan, Michael
  Submitted to: North American Alfalfa Improvement Conference
                                    Publication Type: Abstract Only
                                    Publication Acceptance Date: 6/6/2006
                                    Publication  Date: 6/16/2006
                                    Citation: Sullivan, M.L. 2006. Optimizing efficiency and flexibility of agrobacterium-mediated transformation of red clover (Trifolium pratense). In: Proceeding of the 40th North American Alfalfa Improvement Conference and 19th Trifolium Conference, July 16-19, 2006, Bloomington, MN. Available: http://www.naaic.org/Meetings/National/2006meeting/procedings/Sullivan(poster).PDFInterpretive Summary:
                                Technical Abstract: Genetic modification of plants by the insertion of transgenes can be a powerful experimental approach to answer basic questions about gene product function. This technology can also be used to make improved crop varieties for use in the field. To apply this powerful tool to red clover (Trifolium pratense), germplasm with increased frequency of regeneration in tissue culture was developed (Quesenberry and Smith, 1993, Crop. Sci. 33:585) and utilized as the explant source for development of an Agrobacterium-mediated transformation protocol (Quesenberry et al., 1996, Crop Sci. 36:1045; see http://www.dfrc.ars.usda.gov/DFRCWebPDFs/2006-31_Sull_369_384.pdf for a detailed protocol). We have been testing several variables within this original protocol to increase the efficiency and flexibility of the red clover transformation system. These tests include the use of hygromycin resistance as the plant selectable marker, increasing the duration of Agrobacteria-explant co-cultivation, and comparing A. tumefaciens strains LBA4404 (commonly used in transformation protocols) and EHA105, a kanamycin sensitive derivative of the EHA101 strain used in the original protocol. Hygromycin B at 25 mg/L was effective at selecting for successful transformation of binary vectors containing a hygromycin resistance gene (hpt) under the control of a Nos or double CaMV 35S promoter (Fig. 1). Developing embryos and fully regenerated red clover plants showed expression of a linked GUS transgene with few escapes. Hygromycin B was also highly effective at suppressing a fungal contaminant we often see on explants derived from greenhouse grown plant material. Preliminary data from experiments using A. tumefaciens strain EHA101 suggest that increasing Agrobacteria-explant co-cultivation time from 2 to 4 days results in a 30-50 % increase in transformation efficiency as assessed by embryo formation on selective media. Comparison of transformation efficiency of A. tumefaciens strains LBA4404 and EHA105 is currently under way. Preliminary results suggest that for red clover there is little difference in transformation efficiency between these two strains. These findings expand the options available to researchers wishing to transform red clover.


Return to top


Connect with ARS Sign up


ARS Home
                            USDA.gov
                            Plain Writing
                            Policies & Links
                            Civil Rights Statements
                            FOIA
                            Accessibility Statement
                            Privacy Policy
                            Non-Discrimination Statement
                            Quality of Information
                            USA.gov
                            WhiteHouse.gov