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| Status |
Public on Apr 23, 2013 |
| Title |
control CO |
| Sample type |
SRA |
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| Source name |
Zhongzhu 1
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| Organism |
Boehmeria nivea |
| Characteristics |
cultivar: Gaud
tissue: leaf, root, stem and shoot
developmental stage: 30-day-old
treatment: drought stress
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| Treatment protocol |
the potted 30-day-old ramies were transferred to a movable rain-off shelter and were parted as two groups. One group (CO) was used as control by watering daily and the other group (DS) was treated for drought stress by withholding water. Each group was planted with three replicates. The degree of drought stress in DS ramie was determined by monitoring the relative water contents (RWC) of the ramie leaves.
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| Extracted molecule |
total RNA |
| Extraction protocol |
After severe drought stress, The CO and DS ramie tissues samples of three replicates including leaf, root, stem and shoot were individually collected. The sampled tissues were immediately frozen in liquid nitrogen and stored at -80° until use. The same tissue sample of three replicates of each group was mixed to extract RNA. Total RNAs of two treatment ramie were extracted from each tissue sample using TRIzol reagent (Transgene Company, Illkirch Graffenstaden Cedex, France) according to the manufacturer’s protocol. The RNA was equally pooled from the five tissues for cDNA library preparation
Six micrograms of total RNA for CO and DS ramie was individually purified using biotin-Oligo (dT) magnetic bead adsorption. First- and second-strand cDNA synthesis was performed after the RNA was bound to the beads. While on the beads, double strand cDNA was digested with NlaIII endonuclease to produce a bead-bound cDNA fragment containing sequence from the 3’-most CATG to the poly (A)-tail. These 3’ cDNA fragments were purified using magnetic bead precipitation and the Illumina adapter 1 (GEX adapter 1) was added to new 5’ end. The junction of Illumina adapter 1 and CATG site was recognized by MmeI, which is a Type I endonuclease (with separated recognition sites and digestion sites). The enzyme cuts 17 bp downstream of the CATG site, producing 17 bp cDNA sequence tags with adapter 1. After removing 3’ fragments with magnetic bead precipitation, the Illumina adapter 2 (GEX adapter 2) was ligated to 3’ end of the cDNA tag. These cDNA fragments represented the tag library.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
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| Data processing |
Tag sequencing: The 3'-tag molecules were fixed onto the Illumina Sequencing Chip (flow cell). Each molecule grew into a single-molecule cluster sequencing template through in situ amplification. Four color-labeled nucleotides were added, and sequencing was performed with the method of sequencing by synthesis. Image analysis and base calling were performed using the Illumina Pipeline, and cDNA sequence tags were revealed after purity filtering. The tags passing initial quality tests were sorted and counted. Each tunnel generates millions of raw reads with sequencing length of 49 bp (target tags plus 3’adaptor). Each molecule in the library represented a single tag derived from a single transcript.
Filtering: low quality tags such as tags containing ‘N’ and adaptor sequences were filtered
Gene annotation :“Clean Tags” were obtained by filtering off adaptor-only tags and low-quality tags (containing ambiguous bases). Comparison of the Clean Tags sequences with ramie transcriptome sequence (published in BMC genomics, Liu et al. 14:125) by BLASTN was carried out. All clean tags were annotated based on ramie reference genes. The number of annotated clean tags for each gene was calculated and then normalized to TPM (number of transcripts per million clean tags)
Genome_build: ramie transcriptome sequence (SRA057664)
Supplementary_files_format_and_content: the text file contain the tag sequence, copy number and TPM value
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| Submission date |
Apr 22, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Touming Liu |
| E-mail(s) |
liutouming@caas.cn
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| Phone |
86 731 88998580
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| Organization name |
Chinese Academy of Agricultural Sciences
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| Department |
Institute of Bast Fiber Crops
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| Street address |
NO.348, western road of xianjia lake
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| City |
Changsha |
| ZIP/Postal code |
410205 |
| Country |
China |
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| Platform ID |
GPL17045 |
| Series (1) |
| GSE46253 |
Identification of drought stress-responsive transcription factor in ramie (Boehmeria nivea L．Gaud) |
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| Relations |
| BioSample |
SAMN02054415 |
| SRA |
SRX270939 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1127036_CO_TagCopyNumber.txt.gz |
2.0 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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