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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Dec 07, 2019 |
| Title |
A2 |
| Sample type |
SRA |
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| Source name |
fruit
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| Organism |
Cucumis sativus |
| Characteristics |
tissue: Ventral side of fruit
age: 2 days after anthesis
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from tissue or cell samples by using TRIzol Reagent (Ambion) following the manufacturer’s instructions. RQ1 DNase (promega) was added to remove DNA. The quality and quantity of the purified RNA were determined by measuring the absorbance at 260 nm/280 nm (A260/A280) using smartspec plus (BioRad). RNA integrity was further verified by 1.5% Agarose gel electrophoresis.
For each sample, 10 μg of total RNA was used for RNA-seq library preparation. Polyadenylated mRNAs were purified and concentrated with oligo (dT)-conjugated magnetic beads (invitrogen) before used for directional RNA-seq library preparation. Purified mRNAs were ion fragmented at 95 ℃ followed by end repair and 5' adaptor ligation. Then mRNA reverse transcription was performed with RT primer harboring 3' adaptor sequence and randomized hexamer. The cDNAs were purified and amplified. And products corresponding to 200-500 bps were purified, quantified and stored at -80 ℃ before sequencing. Illumina HiSeq 2000 platform was used to generating paired-end reads. Illumina GAIIx platform was used to generating single-end reads.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina Genome Analyzer IIx |
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| Data processing |
Raw reads were first discarded if containing more than 2-N bases, then reads were processed by clipping adaptor, removing low quality bases, and discarding too short reads (less than 16nt). FASTX-Toolkit (Version 0.0.13) was used to get the clean reads.
After that, clean reads were aligned to the Cucumber v2.0 genome by tophat2. Aligned reads with more than one genome location were discarded due to their ambiguous location.
http://cmb.bnu.edu.cn/Cucumis_sativus_v20/. Download Cucumis_sativus_v20-Cucumber_v2.cds.fasta file. For example, to verify the Csa1G014390 gene, change the G into M, query this gene. (G is representative of the gene, M represents the mRNA, you download the CDS sequence, so it is M, not G. When we analyze sequence data, download the genome sequence, so expressed gene_value is G.)
Uniquely localized reads were used to calculate reads number and RPKM value (RPKM represents reads per kilobase and per million) for each gene according to reads and genes genome location. Other statistical results, such as gene coverage and depth, reads distribution around start codon and stop codon, were also obtained.
After getting the Expression level of all genes in all samples, differentially expressed genes were analyzed by using edgeR(Robinson et al., 2010), one of R packages. 0.01 p-value and 2 fold change were set as the threshold to define DEGs.
Genome_build: C.sativus V2.0
Supplementary_files_format_and_content: tab-delimited MS Excel file including RPKM values for each Sample
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| Submission date |
Dec 09, 2016 |
| Last update date |
Dec 07, 2019 |
| Contact name |
chunhua wang |
| E-mail(s) |
haowch@163.com
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| Phone |
18246196785
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| Organization name |
Notheast Agriculture Unversity
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| Department |
Horticulture
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| Lab |
Lab of Cucumber Molecular breeding
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| Street address |
Mucai Street 59
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| City |
harbin |
| State/province |
Heilongjaing |
| ZIP/Postal code |
150030 |
| Country |
China |
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| Platform ID |
GPL22771 |
| Series (1) |
| GSE91085 |
Transcriptome Profiling of bending fruit and Functional Analysis of an ERF/AP2 Family Gene CsERF025 Involved in bending fruit development in cucumber (Cucumis sativus L.) |
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| Relations |
| BioSample |
SAMN06128000 |
| SRA |
SRX2408997 |
| 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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