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| Status |
Public on Mar 16, 2019 |
| Title |
LT_YFB_S_R3 |
| Sample type |
SRA |
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| Source name |
young fruiting body, stipe
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| Organism |
Lentinus tigrinus |
| Characteristics |
strain: RLP-9953-sp
group: LT_YFB_S
flowcell_id: C86WFANXX
tissue: young fruiting body, stipe
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| Growth protocol |
Vegetative mycelia of Lentinus tigrinus RLP-9953-sp were maintained on MEA (20 g malt extract, 0.5 g yeast extract, 15 g agar for 1L). For fruiting a mycelial plug was placed (5 mm diameter) on modified sawdust-rice bran medium (1 part wheat bran and 2 parts aspen sawdust wetted to 65% moisture for 100 ml in a 250 ml beaker). The culture was incubated for 21 days at 30 °C in the dark, then placed in a moist growth chamber at 25 °C in a 12/12 hour light/dark cycle. Vegetative mycelia, stage 1 primordia, stage 2 primordia cap and stipe, young fruiting body cap and stipe and fruiting body cap and stipe tissues were harvested for RNA-Seq. Stage 1 primordium was defined as a 5-20 mm tall white stalk-structure without any differentiation of a cap initial, stage 2 primordium was defined as a 15 – 25 mm tall stalk-like structure with a brown apical pigmentation (cap initial), young fruiting body had and up to 5 mm wide brown cap initial with just barely visible gills on the bottom, growing on a 30-40 mm tall stipe, fruiting body was 50-70 mm tall, with fully flattened (but not funnel-shaped) cap.
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
After harvesting, samples were immediately placed on liquid nitrogen and stored at -80 °C until use. 50-75 mg frozen tissue was placed in a pre-chilled mortar and ground to a fine powder. RNeasy Midi Kit (QIAGEN) was used for RNA extraction according to manufacturer«s instructions.
Whole transcriptome sequencing was performed using the TrueSeq RNA Library Preparation Kit v2 (Illumina) according to the manufacturer’s instructions. Briefly, RNA quality and quantity measurements were performed using RNA ScreenTape and Reagents on TapeStation (all from Agilent) and Qubit (ThermoFisher); only high quality (RIN >8.0) total RNA samples were processed. Next, RNA was DNaseI (ThermoFisher) treated and the mRNA was purified and fragmented. First strand cDNA synthesis was performed using SuperScript II (ThermoFisher) followed by second strand cDNA synthesis, end repair, 3’-end adenylation, adapter ligation and PCR amplification. All of the purification steps were performed using AmPureXP Beads (Backman Coulter). Final libraries were quality checked using D1000 ScreenTape and Reagents on TapeStation (all from Agilent). Concentration of each library was determined using either the QPCR Quantification Kit for Illumina (Agilent) or the KAPA Library Quantification Kit for Illumina (KAPA Biosystems)
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
Reads were quality trimmed using CLC Genomics Workbench Tool (v 9.5.2). Ambiguous base limit=0, Error probability cutoff value=0.05.
Reads were mapped using RNA-Seq analysis package v 2.1 of CLC Genomics Workbench Tool (v.9.5.2) allowing intergenic mapping. Parameters: min. read length fraction = 0.8, min. read similarity fraction = 0.8. mismatch cost = 2, insertion cost = 3, deletion cost = 3, Strand specific = both, Maximum number of hits for a read=30, Count paired reads as two=no, Expression value= Total counts.
Genes were filtered based on their expression levels in R (v. 3.0.2) keeping only those features that were detected by at least 5 mapped reads in at least 25% of the samples included in the study.
Trimmed mean of M values (TMM) scale normalization was used. Function: calcNormFactors, Package: edgeR, Version 3.4.2
Voom log2 transformation together with quantile normalization was applied. Function: voom normalize=quantile, Package: limma, Version: 3.18.13
Linear modeling with empirical Bayes moderation was built in limma. Package: limma, Version: 3.18.13
Genome_build: Lentinus tigrinus ALCF2SS1-6 v1.0 (Lenti6_1) JGI project ID: 1020066
Supplementary_files_format_and_content: Comma separated files containing RNA-Seq count data, exported from CLC Genomics Workbench Tool
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| Submission date |
Jan 16, 2019 |
| Last update date |
Mar 17, 2019 |
| Contact name |
Laszlo G. Nagy |
| E-mail(s) |
lnagy@fungenomelab.com
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| Phone |
003662599600
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| Organization name |
Biological Research Centre, Hungarian Academy of Sciences
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| Department |
Institute of Biochemistry, Synthetic and Systems Biology Unit
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| Lab |
Laboratory of Fungal Genomics and Evolution
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| Street address |
Temesvari krt 62
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| City |
Szeged |
| State/province |
Csongrad |
| ZIP/Postal code |
6726 |
| Country |
Hungary |
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| Platform ID |
GPL26054 |
| Series (2) |
| GSE125190 |
Transcriptomic atlas of mushroom development reveals conserved genes behind complex multicellularity in fungi [Lentinus tigrinus] |
| GSE125200 |
Transcriptomic atlas of mushroom development reveals conserved genes behind complex multicellularity in fungi |
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| Relations |
| BioSample |
SAMN10756312 |
| SRA |
SRX5255207 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM3564904_LT_YFB_S_R3.CLC_RNA-Seq.csv.gz |
369.4 Kb |
(ftp)(http) |
CSV |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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