NCBI Logo
GEO Logo
   NCBI > GEO > Accession DisplayHelp Not logged in | LoginHelp
GEO help: Mouse over screen elements for information.
          Go
Sample GSM1325467 Query DataSets for GSM1325467
Status Public on Jun 30, 2014
Title IPO323_Ta
Sample type SRA
 
Source name infected wheat leaf
Organism Zymoseptoria tritici
Characteristics strain: IPO323
tissue: leaf
age: 4 days post inoculation
genotype: wild type
Growth protocol Cultures were inoculated onto solid YMS agar (4 g yeast extract, 4 g malt extract, 4 g sucrose, 20 g bacto agar, 1 l H2O) at 18°C. For transcriptional profiling of axenic culture, yeast-like cells were isolated from these plates after 72 h. For transcriptional profiling of plant infections, 15 day old wheat seedlings of the cultivar Obelisk and the Brachypodium distachyon accession Bd21 were inoculated as follows. A distinct area of the second leaf (10 - 15 cm) was marked and a spore solution of 1 x 107 yeast cells/ml containing 0.1% Tween 20 was brushed onto these areas. After an initial incubation for 48 h at 100% humidity and 22°C, inoculated plants were incubated with a 16 h light period at 75% humidity and 22°C for four days.
Extracted molecule total RNA
Extraction protocol To extract total RNA from fungal axenic culture or freeze-dried leaf tissue infected with Z. tritici, 100 mg of sample was crushed in liquid nitrogen and total RNA was extracted using the TRIZOL reagent (Invitrogen) following the manufacturer’s protocol. RNA samples were purified twice with Dynal oligo(dT) magnetic beads (Invitrogen), following the manufacturer's instructions.
RNA-seq libraries for plant infection replicates were prepared from an input of 4 µg total purified polyA RNA according to recommendations of the supplier (TruSeq RNA sample preparation v2 guide, Illumina).
 
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 2000
 
Data processing Quality evaluation using FastQC
Trimming: 12 bp from left and 5 bp from right for: 234_A, 234_B, 234_C, 234_D, 234_E, 234_F
Quality filtering and Masking: Reads with an overall quality score below 20 as well as reads having less than 25% of nucleotides with a quality score above 20 were discarded. For the remaining reads nucleotides with quality scores below 20 were masked with Ns.
Filtering for plant reads: In order to account for incorrect mapping of plant reads with high similarity to fungal sequences, reads from infected plant tissue were mapped to both the plant genomes and the genome of Z. tritici IPO323. All plant reads were removed for further downstream analyses. To this end reads with 100% identity to the plant genomes and a maximum of 25% N positions were filtered out in fastq_screen v0.4.1 (www.bioinformatics.babraham.ac.uk/projects/fastq_screen) applying a sensitive local Bowtie2 alignment
Read Mapping: Mapping of single end RNAseq reads to the genome of the Z. tritici isolate IPO323 (Goodwin et al. 2011) was carried out with tophat v2.0.9 (Kim et al. 2013). Mapping was disabled for novel splice discovery and used the JGI transcript annotation of IPO323 as reference (Goodwin et al. 2011; http://genome.jgi.doe.gov/Mycgr3/Mycgr3.home.html). The maximum number of read mismatches and read gap length were set to 2. Reads that mapped to more than 10 locations were eliminated from the analysis. Relative abundances for predicted transcripts of IPO323 were estimated in cufflinks v2.1.1 (Trapnell et al. 2012). Total read counts per transcript were estimated in htseq-count (http://www-huber.embl.de/users/anders/HTSeq/doc/count.html) applying the intersection-strict mode.
Supplementary_files_format_and_content: csv files with FPKM values and total counts for each transcript of IPO323 (Z. tritici)
 
Submission date Feb 11, 2014
Last update date May 15, 2019
Contact name Ronny Kellner
E-mail(s) ronny.kellner@mpi-marburg.mpg.de
Organization name MPI for Terrestrial Microbiology
Lab Fungal Biodiversity Group
Street address Karl-von-Frischstrasse 10
City Marburg
ZIP/Postal code 35043
Country Germany
 
Platform ID GPL18273
Series (1)
GSE54874 Transcriptome sequencing at early stage infection of the wheat pathogen Zymoseptoria tritici reveals chromosomal differences in transcription patterns and host specific gene expression.
Relations
BioSample SAMN02640197
SRA SRX469418

Supplementary file Size Download File type/resource
GSM1325467_234_A.txt.gz 335.2 Kb (ftp)(http) TXT
GSM1325467_234_B.txt.gz 339.3 Kb (ftp)(http) TXT
GSM1325467_234_C.txt.gz 336.5 Kb (ftp)(http) TXT
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

| NLM | NIH | GEO Help | Disclaimer | Accessibility |
NCBI Home NCBI Search NCBI SiteMap