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| Status |
Public on Aug 18, 2014 |
| Title |
NC |
| Sample type |
SRA |
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| Source name |
folded leaf, fully expanded leaf and roots
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| Organism |
Manihot esculenta |
| Characteristics |
tissue: folded leaf, fully expanded leaf and roots
cultivar: SC124
harvest time: 6h, 24h and 5d
treatment: normal condition (NC) of 24°C were watered once with Hoagland’s solution every 5 days
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| Growth protocol |
Stem segments with three nodes of Cassava (Manihot esculenta Crantz) cultivars were extracted from 8-month-old plants, and inclined in 3-L pots filled with barren red soil: vermiculite (1:1, v/v), fertilized with Hoagland’s solution (Hoagland and Arnon 1950), to propagate and generate well-balanced seedlings. The solution was renewed with 300ml quarter-strength solution once a week. After 2 months of planting, the uniform seedlings were subjected to cold stress treatment. All plants were field grown in Haikou, Hainan, China, during April and June of natural conditions (11h light, 13h dark and 25°C during the day and 18°C at night). Cassava cultivar SC124 was transferred to normal 24°C illumination incubator (SANYO, Japan) for 2 days to set a homogenization starting point, and then was subjected to three types of cold treatments. 1) Gradual cold acclimation (CA in Figure S1): temperature was decreased from 24°C to 14°C with the rate of -2°C/h to induce mild cold stress. Temperature was then held constant at 14°C for five days to accommodate cold acclimation. RNA was collected at 6h, 24h and 5d after the temperature reaching 14°C. 2) Cold stress after cold acclimation (CCA): after 5 days of cold acclimation and growth under 14°C, plants were watered once with Hoagland’s solution, transferred further to 4°C by -2°C/h gradient cooling, and cultivated at constant 4°C for another 5d. 3) Cold shock (CS): plants grown under 25°C were subjected to dramatic temperature decline to 4°C with a rate of -4°C/h to ensure the temperature reached 4°C at the same time as the CCA treatment. In the two latter treatments, RNA was collected at 6h, 24h and 5d after temperature reaching 4°C. In parallel, plants grown under the normal condition (NC) of 24°C were watered once with Hoagland’s solution every 5 days, and RNA was collected at 0d, 5d and 10d. The mixture samples of SC124 (details were given below) were subjected to small-RNA and mRNA expression profiling by NextGen deep sequencing.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Three organs/tissues (folded leaf, fully expanded leaf and roots) of Cassava cultivar SC124 harvested at 6h, 24h and 5d for three cold treatments of CA, CCA and CS, for gene expression profiling at the stages of initial response, secondary response, and functional adaption to cold stresses. Total RNA of each sample was isolated individually, and then pooled with an equal amount from each sample into one for profiling. As a result, four mRNA libraries and four small-RNA libraries, corresponding to the conditions of CA, CCA, CS and NC, were constructed.
The four mRNA libraries were sequenced by RNA-seq by Illumina GAII following Illumina RNA-seq protocol. Briefly, total RNAs were isolated, purified and reversely transcribed, the resulting cDNA products were subsequently digested with NlaIII and the 3′-cDNA fragments captured with the oligo(dT) beads, and then ligated to the Illumina GEX NlaIII Adapter 1. The junction of Illumina adapter 1 and CATG site contained the recognition site of MmeI, which cutting 17 bp downstream of the recognition site (CATG) to produce tags. After removing 3′fragments with magnetic beads precipitation and MmeI digestion, an Illumina GEX adapter 2 was introduced at the end of tags. The resulting adapter-ligated cDNA tags were subjected to 15 cycles of linear PCR amplified, purified and sequenced with the method of sequencing by synthesis (SBS) using Illumina GAII
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina Genome Analyzer IIx |
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| Description |
normal condition (NC) of 24°C were watered once with Hoagland’s solution every 5 days
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| Data processing |
Raw RNA-Seq reads with low quality were firstly discarded. Sequencing adaptors were trimmed using an in-house method that recursively searches for the longest substring of the adaptor appearing within a sequence read. If a raw sequence read did not have a substring of the adaptor longer than 6nt, it was considered as having no adaptor.
The adaptor-trimmed sequences with no ambiguous reads, which were referred to as qualified reads, were then mapped to the cDNA sequences of Cassava using Bowtie 0.12.7 (Langmead et al. 2009) allowing no more than one mismatch.
Reads Count Per Million (CPM) were calculated for each transcript
Raw count of reads were also normalized using upper-quatertile normalization method for differential expression analysis
Genome_build: n/a
Supplementary_files_format_and_content: tab-delimited text files include CPM values and upper-quartile normalized counts for each sample (PACid from Phytozome and Mesculenta_147_PfamAB.txt rom comailab.genomecenter.ucdavis.edu)
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| Submission date |
Nov 07, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Weixiong Zhang |
| E-mail(s) |
[email protected]
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| Organization name |
Washington university in St Louis
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| Street address |
Campus Box 1045 One Brookings Drive
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| City |
Saint Louis |
| State/province |
MO |
| ZIP/Postal code |
63130 |
| Country |
USA |
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| Platform ID |
GPL13617 |
| Series (2) |
| GSE52176 |
Moderate stress acclimation provides immunity to stress by rewiring regulatory networks and inducing genes with protective functions in Cassava |
| GSE52178 |
Stress acclimation in Cassava |
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| Relations |
| BioSample |
SAMN02400134 |
| SRA |
SRX374719 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1260850_NC.cpm.tsv.gz |
276.8 Kb |
(ftp)(http) |
TSV |
| GSM1260850_NC.qnorm.tsv.gz |
260.5 Kb |
(ftp)(http) |
TSV |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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