A high throughput system for transposon tagging and promoter trapping in tomato

A high throughput system for transposon tagging and promoter trapping in tomato We describe new tools for functional analysis of the tomato genome based on insertional mutagenesis with the maize Ac/Ds transposable elements in the background of the miniature cultivar Micro‐Tom. 2932 F3 families, in which Ds elements transposed and were stabilized, were screened for phenotypic mutations. Out of 10 families that had a clear mutant phenotype, only one mutant was Ds‐tagged. In addition, we developed promoter trapping using the firefly luciferase reporter gene and enhancer trapping, using β‐glucuronidase (GUS). We show that luciferase can be used as a non‐invasive reporter to identify, isolate and regenerate somatic sectors, to study the time course of mutant expression, and to identify inducible genes. Out of 108 families screened for luciferase activity 55% showed expression in the flower, 11% in the fruit and 4% in seedlings, suggesting a high rate of Ds insertion into genes. Preferential insertion into genes was supported by the analysis of Ds flanking sequences: 28 out of 50 sequenced Ds insertion sites were similar to known genes or to ESTs. In summary, the 2932 lines described here contain 2–3 Ds inserts per line, representing a collection of approximately 7500 Ds insertions. This collection has potential for use in high‐throughput functional analysis of genes and promoter isolation in tomato. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png The Plant Journal Wiley

A high throughput system for transposon tagging and promoter trapping in tomato

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Abstract

We describe new tools for functional analysis of the tomato genome based on insertional mutagenesis with the maize Ac/Ds transposable elements in the background of the miniature cultivar Micro‐Tom. 2932 F3 families, in which Ds elements transposed and were stabilized, were screened for phenotypic mutations. Out of 10 families that had a clear mutant phenotype, only one mutant was Ds‐tagged. In addition, we developed promoter trapping using the firefly luciferase reporter gene and enhancer trapping, using β‐glucuronidase (GUS). We show that luciferase can be used as a non‐invasive reporter to identify, isolate and regenerate somatic sectors, to study the time course of mutant expression, and to identify inducible genes. Out of 108 families screened for luciferase activity 55% showed expression in the flower, 11% in the fruit and 4% in seedlings, suggesting a high rate of Ds insertion into genes. Preferential insertion into genes was supported by the analysis of Ds flanking sequences: 28 out of 50 sequenced Ds insertion sites were similar to known genes or to ESTs. In summary, the 2932 lines described here contain 2–3 Ds inserts per line, representing a collection of approximately 7500 Ds insertions. This collection has potential for use in high‐throughput functional analysis of genes and promoter isolation in tomato.

Journal

The Plant JournalWiley

Published: May 1, 2000

References

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