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Status |
Public on Aug 08, 2019 |
Title |
luh4_4h |
Sample type |
SRA |
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Source name |
roots_with Al
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Organism |
Arabidopsis thaliana |
Characteristics |
tissue: root old of seedlings: six-days time point: 4h genotype/variation: luh-4 mutant treament: with Al ecotype: Columbia
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Treatment protocol |
Approximately 500 seedlings (6-d-old) of both the wild-type Col and luh-4 mutant line were exposed to a nutrient solution containing 0 or 25 μM AlCl3 (pH4.2). After 4 h, the roots were sampled for RNA isolation.
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Growth protocol |
The seedlings were pre-grown in nutrient solution for 6 d at pH5.5 in a growth chamber with a 16/8 h light/dark cycle at 22 oC.
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Extracted molecule |
total RNA |
Extraction protocol |
Total RNA was isolated using the RNeasy® Plant Mini Kit (QIAGEN GmbH, Qiagen, GmbH, Hilden, Germany) following the manufacturer’s protocol. RNA libraries were prepared for sequencing using standard Illumina protocols The total RNA samples are first treated with DNase I to degrade any possible DNA contamination. Then the mRNA is enriched by using the oligo(dT) magnetic beads (for eukaryotes). Mixed with the fragmentation buffer, the mRNA is fragmented into short fragments (about 200 bp). Then the first strand of cDNA is synthesized by using random hexamer-primer. Buffer, dNTPs, RNase H and DNA polymerase I are added to synthesize the second strand. The double strand cDNA is purified with magnetic beads. End reparation and 3'-end single nucleotide A (adenine) addition is then performed. Finally, sequencing adaptors are ligated to the fragments. The fragments are enriched by PCR amplification. During the QC step, Agilent 2100 Bioanaylzer and ABI StepOnePlus Real-Time PCR System are used to qualify and quantify of the sample library. The library products are ready for sequencing via Illumina HiSeqTM 2000 or other sequencer when necessary.
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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 |
By base calling, the original image data produced by the sequencer is transferred into sequences, which are defined as "raw reads"(or "raw data") and saved as ".fastq" files. As the raw reads may contain low quality reads and or adaptor sequences, preprocessing is necessary before starting further analysis. Normally, data cleaning (or data filtering) is performed to obtain "clean reads" (or "clean data") for further analysis. As there are some adaptor sequences and/or low quality reads present in the raw reads, data filtering is carried out to obtain high quality reads as the clean reads (clean data). The procedure include following steps: 1. Remove reads with adaptor sequences. 2. Remove reads in which the percentage of unknown bases (N) is greater than 10%. 3. Remove low quality reads. If the percentage of the low quality base (base with quality value ≤ 5) is greater than 50% in a read, we define this read as low quality The gene expression level is calculated by using RPKM[Mortazavi et al. (2008). Nature Methods. 5(7): 621-8] method (Reads Per kb per Million reads), Clean reads were mapped to reference sequences and/or reference gene set using SOAPaligner/SOAP2[Li R et al. (2009) Bioinformatics, 25 (15). 1966-1967.]. No more than 2 mismatches were allowed in the alignment Genome_build: TAIR10 Supplementary_files_format_and_content: Excel files include RPKM values for each Sample
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Submission date |
Aug 08, 2016 |
Last update date |
Aug 08, 2019 |
Contact name |
Zhong-Bao Yang |
E-mail(s) |
zbyang@sdu.edu.cn
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Organization name |
Shandong University (Qingdao)
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Department |
School of Life Science
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Lab |
he Key Laboratory of Plant Development and Environmental Adaptation Biology, Ministry of Education
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Street address |
72# Binhai Road
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City |
Qingdao |
State/province |
Shandong |
ZIP/Postal code |
266237 |
Country |
China |
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Platform ID |
GPL13222 |
Series (1) |
GSE85292 |
The luh-4 mutant has a differential transcriptional program in response to Al stress |
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Relations |
BioSample |
SAMN05521267 |
SRA |
SRX2007316 |
Supplementary file |
Size |
Download |
File type/resource |
GSM2264187_luh4_4h.Gene.rpkm.txt.gz |
1.9 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
Raw data are available in SRA |
Processed data provided as supplementary file |
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