Identification of direct target genes of the tomato fruit-ripening regulator RIN by promoter ChIP-chip

The tomato (Solanum lycopersicum) MADS-box transcription factor RIN, one of the earliest-acting fruit-ripening regulators, is required for both ethylene-dependent and -independent regulatory pathways for fruit ripeing. Here, we performed chromatin immunoprecipitation coupled with a DNA microarray (ChIP-chip) for the putative promoters of whole tomato predicted genes (ITAG2) for the genome-wide identification of the direct RIN targets. The ChIP-chip with anti-RIN antibody resulted in detection of 1,046 RIN-binding sites, each of which was assigned a significantly high peak score (FDR≤0.05) in at least two of the three biologically independent analyses. Using the information about genomic position of the RIN-binding sites, we found 1,200 genes as potential direct RIN targets that carried one or more RIN-binding sites in the transcription regulatory region (2-kb upstream putative promoter) or in other gene regions, such as exons, introns or a downstream region from the translation termination site (1-kb), where the promoter region of a neighbor gene are overlapped. Three biologically independent samples (chromatin-immunoprecipitated DNA and input DNA) recovered from the ripening tomato fruits.