Comment[ArrayExpressAccession] E-GEOD-49289 MAGE-TAB Version 1.1 Public Release Date 2014-06-23 Investigation Title Transcriptome profiling of tomato fruits of a FRUITFULL-suppressed line and rin mutant by next generation RNA sequencing Comment[Submitted Name] Transcriptome profiling of tomato fruits of a FRUITFULL-suppressed line and rin mutant by next generation RNA sequencing Experiment Description The tomato MADS-box FRUITFULL (FUL) homologs, FUL1 and FUL2, interact with the main ripening regulator RIPENING INHIBITOR (RIN). To clarify their role in fruit ripening, we generated FUL1/FUL2-suppressed transgenic lines by RNAi. We found that five transgenic lines bearing fruits that did not ripen normally: lycopene accumulation and increase of ethylene production were severely inhibited. We then performed next generation RNA sequencing (RNA-Seq) analysis of the fruits of a FUL1/FUL2-suppressed line (TF18) with those of the wild type (Ailsa Craig cultivar; AC) and rin mutant. The comparison of RNA-Seq data among them indicated that FUL1/FUL2-suppression significantly affected the expression of a larger portion of ripening-induced and -repressed genes than the rin mutation did. Moreover, the effect of FUL1/FUL2-suppression was observed not only in the fruits harvested at the wild type ripening age [45 days after pollination (DAP)] but also in those at the pre-ripening age (35 DAP). This suggests that the FUL homologs play an essential role in the regulation of fruit development and ripening, the role which covers a wider range of biological processes than RIN does. Differentially expressed genes (DEGs) between the wild type and TF18 fruits included known ripening-related genes such as ACS2 and ACS4 involved in ethylene production and PSY1 in carotenoid biosynthesis, consistent with the phenotype of TF18 fruits described above. The DEGs also included many direct RIN target genes, which supports the hypothesis that the FUL homologs regulate fruit ripening in a form of MADS-box complex with RIN. mRNA profiles of wild type (Ailsa Craig cultivar), rin mutant and FUL1/FUL2-suppressed tomato fruits harvested at 35DAP and 45 DAP were generated by next generation sequencing, in triplicate, using Illumina Hiseq2000. Term Source Name ArrayExpress EFO Term Source File http://www.ebi.ac.uk/arrayexpress/ http://www.ebi.ac.uk/efo/efo.owl Person Last Name Ito Fujisawa Shima Nakano Ito Person First Name Yasuhiro Masaki Yoko Toshitsugu Yasuhiro Person Email yasuito@affrc.go.jp Person Affiliation National Food Research Institute Person Address Food Biotechnology Division, National Food Research Institute, 2-1-12 Kannondai, Tsukuba, Ibaraki, Japan Person Roles submitter Protocol Name P-GSE49289-4 P-GSE49289-1 P-GSE49289-3 P-GSE49289-2 Protocol Description Illumina CASAVA software (ver.1.8.1) was used for basecalling. Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to the predicted protein-coding sequences (CDS) of tomato genes provided the International Tomato Annotation Group (ITAG, version2; available at http://solgenomics.net/organism/Solanum_lycopersicum/genome) using bowtie2 with parameters -q -p 4. Reads Per Kilobase of exon per Million mapped reads (RPKM) were calculated for all tomato predicted genes. Genome_build: ITAG2_cds Supplementary_files_format_and_content: A tab-delimited text file includes RPKM values for each sample. The harvested fruits were sliced and immediately frozen in liquid nitrogen and stored at -8C. The frozen fruits were powdered with moter and pestle. Total RNA samples were prepared from fruits of wild type, a FUL1/FUL2 suppressed line (TF18) and rin mutant at 35 DAP (G stage) and 45 DAP (P-stage) by SDS-phenol extraction and following elimination of polysaccharides with 2-botoxyethanol. The RNAs were further purified with the RNeasy Plus Kit (Qiagen). The quality of the RNAs was confirmed using a 2100 Bioanalyzer (Agilent). RNA libraries were prepared for sequencing using standard Illumina protocols. Plants were grown in a glasshouse under 16-h light at 26C and 8-h dark at 20C conditions, and the fruits were harvested at 35 and 45 DAP. Protocol Type normalization data transformation protocol sample treatment protocol nucleic acid library construction protocol growth protocol Experimental Factor Name developmental stage genotype Experimental Factor Type developmental stage genotype Publication Title Transcriptional regulation of fruit ripening by tomato FRUITFULL homologs and associated MADS box proteins. Publication Author List Fujisawa M, Shima Y, Nakagawa H, Kitagawa M, Kimbara J, Nakano T, Kasumi T, Ito Y PubMed ID 24415769 Publication DOI 10.1105/tpc.113.119453 Comment[AEExperimentDisplayName] Transcription profiling by high throughput sequencing of tomato of FUL1/FUL2-suppressed fruits harvested at preripening (mature green) stage and ripening (pink coloring) stage Comment[SecondaryAccession] GSE49289 Comment[GEOReleaseDate] 2014-06-23 Comment[ArrayExpressSubmissionDate] 2013-07-27 Comment[GEOLastUpdateDate] 2014-06-23 Comment[AEExperimentType] RNA-seq of coding RNA Comment[AdditionalFile:Data1] GSE49289_Fujisawa_etal_NGS_FULRNAi_RPKM.txt Comment[SecondaryAccession] SRP028278 Comment[SequenceDataURI] http://www.ebi.ac.uk/ena/data/view/SRR943813-SRR943830 SDRF File E-GEOD-49289.sdrf.txt