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| Status |
Public on Nov 24, 2018 |
| Title |
NM3 |
| Sample type |
SRA |
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| Source name |
no molecular signal added_roots
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| Organism |
Triticum aestivum |
| Characteristics |
cultivar: Chinese Spring
treatment: non-mycorrhizal treatment (NM)
tissue: roots
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| Treatment protocol |
Two treatments were established: (1) non-mycorrhizal treatment (NM), in which there was no AM inoculum in pots; and (2) molecular signals communication treatment (MSC), in which each pot contained 15 bags filled with 25 g of the inoculum of Rhizophagus irregularis (BGC-HEB07D). The bags were 10 cm × 10 cm and were made of a 0.25 µm filter membrane. This membrane can physically separate wheat roots, AM spores and AM hyphae but allows the communication of the molecular signals secreted by the host plants or the AM fungus. To eliminate the influence of the buried bags on the growth of wheat, 15 bags of the same size were filled with 20 g of steam-sterilized inoculum of R. irregularis (to maintain a similar microbial community with the bags used in the MSC treatment, the filtrate from the unsterilized AM inoculum was added to the bags) were also buried in the pots of the NM treatments.
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| Growth protocol |
The experiment was conducted in Northwest A&F University, Yangling, Shaanxi province, China (34.26°N, 108.06°E) from 23th March-13th April, 2017. Wheat seeds (T. aestivum L., Cultivar: Chinese spring) were surface-sterilized by submerging with 1% NaClO for 10 min, rinsed with deionized water, and then pre-germinated on moist filter paper in the dark at 25 °C for 24 h. The pre-germinated wheat seeds were planted into pots filled with approximately 2.5 L of steam-sterilized soil-quartz sand mixture (1:3) (v/v). The soil was clay loam and was classified as a Eum-Orthic Anthrosol (FAO Soil Classification, Gong et al. 2003). Its basic fertility characteristics were: nitrate 10.2 mg kg-1, ammonium 1.95 mg kg-1, available phosphorus (P) (Olsen-P) 7.35 mg kg-1, available potassium (K) 143.0 mg kg-1 and organic matter 8.55 g kg-1.Twenty-five pre-germinated wheat seeds were planted into each pot, and plants were thinned to 15 plants for each pot 5 days after seeding. Wheat plants were cultured outdoors and watered when necessary.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from approximately 100 mg of wheat root of three biological replicates of the two treatments using the RNeasy Plant Mini Kit (Qiagen, USA). according to the manufacturer’s instructions.The concentration and quality of the total RNA were estimated using a NanoDrop Spectrometer (Thermo; USA) and verified using agarose gel electrophoresis and an Agilent 2100 bioanalyzer (Agilent, USA).
mRNA of each sample was respectively enriched by Oligo(dT) beads, then the enriched mRNA was fragmented into short fragments (~300bp) using fragmentation buffer, followed by reverse transcription into cDNA with random primers. Second-strand cDNA were synthesized by DNA polymerase I, RNase H, dNTP and buffer. Then the cDNA fragments were purified with QiaQuick PCR extraction kit, end repaired, poly(A) added, and ligated to Illumina sequencing adapters. The ligation products were size selected by agarose gel electrophoresis. Six independent complementary DNA (cDNA) libraries corresponding to the three biological replicates for each treatment were constructed. cDNA library quality and fragment length were verified using an Agilent 2100 bioanalyzer and real-time PCR. Fragment length distributions were within the expected ranges according to the CLC Sequence Viewer software, and the average read length was 150 bp.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina Genome Analyzer |
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| Description |
NM_fpkm
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| Data processing |
Sequences obtained for each of the six libraries were imported and individually processed using RSEM version 1.3.0
For quality filtering, a trimming step was performed on raw reads with the following conditions: (1) removing reads containing adapters; (2) removing reads containing more than 10% of unknown nucleotides; and (3) removing low-quality reads containing more than 50% of low-quality (Q-value ≤ 20) bases. Sequencing coverage was estimated according to the RSEM algorithm by dividing the total number of acquired sequence bases per condition by the number of reference bases. High-quality reads were then mapped against the wheat reference transcriptome “Ensembl release 30 IWGSC1.0” (http://archive.plants.ensembl.org/Triticum_aestivum/Info/Index) using the following mapping parameters: maximum read mismatch, 2; distance between mate-pair reads, 50 bp; and error of distance between mate-pair reads, ±80 bp. To check if there are RNA sequences belonging to R. irregularis, reads were also mapped against the genome of R. irregularis ((http://genome.jgi.doe.gov/Gloin1/Gloin1.home.html) (Tisserant et al. 2013). The reconstruction of transcripts was carried out with software Cufflinks, which, together with TopHat2, allows biologists to identify new genes. All of the reconstructed transcripts were aligned to the reference genome and divided into twelve categories by using Cuffcompare. Genes with class code “u” (the transcripts were unknown or in the intergenic spacer region) were defined as novel genes. The parameters used to identify reliable novel genes were as follows: length of the transcript longer than 200 bp and exon number greater than 2
Genome_build: Ensembl release 30 IWGSC1.0 (http://archive.plants.ensembl.org/Triticum_aestivum/Info/Index)
Supplementary_files_format_and_content: txt file. Gene expression level of each genes
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| Submission date |
Nov 23, 2018 |
| Last update date |
Nov 26, 2018 |
| Contact name |
HUI TIAN |
| E-mail(s) |
tianh@nwsuaf.edu.cn
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| Organization name |
Northwest A&F University
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| Street address |
No.3
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| City |
Yangling |
| ZIP/Postal code |
712100 |
| Country |
China |
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| Platform ID |
GPL10467 |
| Series (1) |
| GSE122851 |
Molecular signal communication in soil during arbuscular mycorrhizal formation changes the transcriptome profiles in wheat (Triticum aestivum L.) roots |
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| Relations |
| BioSample |
SAMN10471542 |
| SRA |
SRX5057643 |
| 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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