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| Status |
Public on Dec 16, 2017 |
| Title |
Shade Pulse, 8 hours since Dawn, Time 0, replicate 1 |
| Sample type |
SRA |
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| Source name |
Clear Day-acclimated culture
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| Organism |
Synechococcus elongatus PCC 7942 = FACHB-805 |
| Characteristics |
hours since light onset (dawn): 8
perturbation: Shade pulse time 0 (prior to perturbation)
light period: 4th
replicate: 1
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| Growth protocol |
Wildtype Synechococcus elongatus PCC 7942 was grown in BG-11M media supplemented with 10 mM HEPES pH 80 constantly bubbled with 1% CO2 in air Cultures were maintained at OD750 03 with periodic dilution Cultures were grown in either a Low Light condition of 50 µmol photons m-2 s-1 or a Clear Day condition where light intensity varied parabolically, reaching a maximum of 600 µmol photons m-2 s-1 Cultures were grown in light for 12 hours, followed by 12 hours of darkness, for two cycles, and then sampled over the third light period On the fourth light period at 8 hours after light onset, we incresed the light intensity of the Low Light culture by 10 fold (High Light pulse) and decreased the light intensity of the Clear Day culture by 10 fold (Shade pulse) for 1 hour prior to returning the cultures to their original culture, and sampled them before, during, and after the perturbation The experiment was repeated twice for 2 biological replicates, and we report both datasets here
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| Extracted molecule |
total RNA |
| Extraction protocol |
25 mL of OD750 03 culture was collected on a cellulose acetate filter and flash frozen Total RNA was isolated from cells using the Qiagen RNeasy mini kit
Ribosomal RNA was depleted from 125 µg of purified RNA using the Illumina Ribo-Zero rRNA removal kit, and strand specific RNA sequencing libraries were prepared using the Illumina Truseq Stranded mRNA sample prep kit
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Description |
Natural_light_RNAseq_processedxlsx
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| Data processing |
Read alignment: Bowtie, v 0129, -v 3 -m 1 -p 16 -t --best --un
Gene expression quantification: alignment output from Bowtie was read into Matlab using custom code To obtain gene expression values, we counted the number of coding-strand sequencing reads that had 5’ ends between the start and stop positions of the coding region of each gene
Gene expression normalization: To normalize gene expression values between samples, we utilized median normalization as described elsewhere (Anders and Huber, 2010) First, we calculated a pseudo-reference for each gene by determining the geometric mean of the expression counts for that gene across all samples, and calculated the ratio of the expression values with the appropriate pseudo-reference value We determined the median value of these ratios within each sample and took this as a size factor that estimates the sequencing depth of each sample To normalize, we divided all gene expression values within a sample by the appropriate size factor Gene expression values were further normalized by dividing the gene expression values by the length of the appropriate open reading frame To easily compare gene expression changes between each condition, we calculated the average expression of a gene over all time points in the Low Light condition Then, we divided the expression of that gene at each time point by the average Low Light expression, and took the Log2 of this ratio
Genome_build: Main chromosome: NC_007604.1; plasmid pANL (plasmid 1), NC_004073.2; plasmid pANS (plasmid 2), KT751091.1
Supplementary_files_format_and_content: Gene expression values for each gene are listed, calculated as described above. Data are reported for two separate biologcal replicates.
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| Submission date |
Sep 25, 2017 |
| Last update date |
May 15, 2019 |
| Contact name |
Joseph Robert Piechura |
| E-mail(s) |
jpiechura25@gmail.com
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| Organization name |
Harvard University
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| Department |
FAS Center for Systems Biology
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| Lab |
Erin O'Shea Lab
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| Street address |
52 Oxford Street
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| City |
Cambridge |
| State/province |
Massachusetts |
| ZIP/Postal code |
02138 |
| Country |
USA |
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| Platform ID |
GPL17750 |
| Series (2) |
| GSE104203 |
Measurement of genome-wide gene expression in cyanobacteria grown under simluated natural light conditions [RNA-Seq] |
| GSE104204 |
Measurement of genome-wide gene expression in cyanobacteria grown under simluated natural light conditions |
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| Relations |
| BioSample |
SAMN07693167 |
| SRA |
SRX3211347 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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