Protocol: fine-tuning of a Chromatin Immunoprecipitation (ChIP) protocol in tomato
2010
Ricardi, Martiniano M | González, Rodrigo M | Iusem, Norberto D
BACKGROUND: Searching thoroughly for plant cis-elements corresponding to transcription factors is worthwhile to reveal novel gene activation cascades. At the same time, a great deal of research is currently focused on epigenetic events in plants. A widely used method serving both purposes is chromatin immunoprecipitation, which was developed for Arabidopsis and other plants but is not yet operational for tomato (Solanum lycopersicum), a model plant species for a group of economically important crops. RESULTS: We developed a chromatin immunoprecipitation protocol suitable for tomato by adjusting the parameters to optimise in vivo crosslinking, purification of nuclei, chromatin extraction, DNA shearing and precipitate analysis using real-time PCR. Results were obtained with two different antibodies, five control loci and two normalisation criteria. CONCLUSION: Here we provide a chromatin immunoprecipitation procedure for tomato leaves that could be combined with high-throughput sequencing to generate a detailed map of epigenetic modifications or genome-wide nucleosome positioning data.
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