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| Status |
Public on Jan 27, 2019 |
| Title |
Mature pollen grains-control conditions (MPGs_C) |
| Sample type |
SRA |
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| Source name |
tomato pollen grains
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| Organism |
Solanum lycopersicum |
| Characteristics |
genotype/cultivar: cv. 3017 (heat-sensitive)
tissue: pollen (male gametophyte)
developmental stage: mature
type: mature pollen grains
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| Growth protocol |
Tomato plants were grown in temperature-controlled greenhouses at the Volcani Center, Bet Dagan, Israel, with day/night temperatures of 26/22 oC, day length of 13–14 h, and under natural illumination conditions until development of the second truss. In separate greenhouses, plants bearing 2-3 inflorescences were exposed to either control conditions (C; maintained at 25 to 26 oC) or to a mild heat treatment (ATT; 1 h at 38 oC followed by 1 h recovery at 25 oC). Plants were watered during the heat treatments to avoid drought stress and wilting.
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| Extracted molecule |
total RNA |
| Extraction protocol |
For transcriptome analysis, pollen grains were derived from plants that were maintained under control conditions and from plants that were exposed to the above-detailed ATT conditions. For each pollen sample, anthers collected from 90 flowers, at developmental stages of three, two, one and zero days before anthesis were dissected and pollen grains were obtained by slicing the anthers transversely and vortexing them in cold, sucrose-free germination solution. The solution was then filtered and pollen grains separated. The released pollen grains were immediately frozen in liquid nitrogen and kept frozen at -80 oC until use. Pollen grains were ground to a fine powder using liquid nitrogen and sea sand (Merck, Darmstadt, Germany) and total RNA was extracted using the Tri-reagent (Sigma Aldrich, Israel). Before further analyses, RNA derived from three stages of pollen maturation (three, two and one days before flower opening, corresponding to post-meiotic immature pollen grains; IPGs) was combined, using equal amounts, into one pool, generating a pooled sample of IPGs RNA, in addition to RNA isolated from mature pollen grains (zero days before flower opening; MPGs), that constituted a separate sample.
For preparation of MACE libraries, poly-adenylated mRNA was isolated from 1 μg of the large fraction of total RNA using Dynabeads® mRNA Purification Kit (Life Technologies GmbH) and cDNA was produced by first and second strand synthesis using the SuperScript® III First-Strand Synthesis System (Life Technologies GmbH). , but with modified, barcoded poly-T adapters that are suitable to bind on the Illumina Hiseq2000 flow cell and that are biotinylated at the very 5′ end. After cDNA production, the biotinylated cDNA was random-fragmented to reach an average size of 250 bps, the 3′ends were captured by streptavidin beads and GenXPro’s TrueQuant adapters were ligated. Ten barcoded samples were sequenced simultaneously in one lane of an Illumina Hiseq2000 with 1 × 94 bps.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
3017PollenA2_MPGs-C
library strategy: MACE (massive analysis of cDNA ends)
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| Data processing |
Sequencing on Illumina HiSeq 2000, basecalling pipeline: HiSeq Control Software 1.5.15.1, RTA 1.13.48.0, CASAVA-1.8.2, Flow cell version: HiSeq Flow Cell v3, Kit version: TruSeq SBS Kit v3
Sequences were cleaned from duplicates ("TrueQuant"), Bases with low sequencing quality were trimmed.
The trimmed reads were mapped to ITAG2.3 Solanum lycopersicum using "SOAP2.2" with the following parameters “-p 16 -r 2 -v 5 -w 10 -g 1“
Expression analysis was performed by in-house scripts and DEGSeq (R-package)
Genome_build: ITAG2.3 Solanum lycopersicum
Supplementary_files_format_and_content: Tab-delimited text files contain read counts, normalized read counts, p-values, fold changes
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| Submission date |
Jul 26, 2018 |
| Last update date |
Jan 27, 2019 |
| Contact name |
GenXPro GenXPro |
| Organization name |
GenXPro GmbH
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| Street address |
Altenhöferallee 3
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| City |
Frankfurt am Main |
| ZIP/Postal code |
60438 |
| Country |
Germany |
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| Platform ID |
GPL16345 |
| Series (1) |
| GSE117733 |
Comparative transcriptome profiling of tomato (Solanum lycopersicum L.) pollen grains during maturation and in response to acquired thermotolerance conditions |
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| Relations |
| BioSample |
SAMN09724763 |
| SRA |
SRX4473904 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3307877_hitCounts_3017PollenA2_PollenAHS.txt.gz |
6.3 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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