Analysis of the chromosomal distribution of transposon-carrying T-DNAs in tomato using the inverse polymerase chain reaction |
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Authors: | Colwyn M Thomas David A Jones James J English Bernard J Carroll Jeffrey L Bennetzen Kate Harrison Alan Burbidge Gerard J Bishop and Jonathan D G Jones |
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Institution: | (1) The Sainsbury Laboratory, John Innes Centre, NR4 7UH Colney Lane, Norwich, UK;(2) Department of Biological Sciences, Purdue University, 47907 West Lafayette, Indiana, USA;(3) Department of Physiology and Environmental Science, University of Nottingham, Faculty of Agricultural and Food Sciences, Sutton Bonnington, Nr. Loughborough, Leicestershire, UK |
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Abstract: | We are developing a system for isolating tomato genes by transposon mutagenesis. In maize and tobacco, the transposon Activator (Ac) transposes preferentially to genetically linked sites. To identify transposons linked to various target genes, we have determined the RFLP map locations of Ac- and Dissociation (Ds)-carrying T-DNAs in a number of transformants. T-DNA flanking sequences were isolated using the inverse polymerase chain reaction (IPCR) and located on the RFLP map of tomato. The authenticity of IPCR reaction products was tested by several criteria including nested primer amplification, DNA sequence analysis and PCR amplification of the corresponding insertion target sequences. We report the RFLP map locations of 37 transposon-carrying T-DNAs. We also report the map locations of nine transposed Ds elements. T-DNAs were identified on all chromosomes except chromosome 6. Our data revealed no apparent chromosomal preference for T-DNA integration events. Lines carrying transposons at known map locations have been established which should prove a useful resource for isolating tomato genes by transposon mutagenesis. |
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Keywords: | Tomato Agrobacterium T-DNA Inverse PCR Activator Transposon tagging |
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