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Sample GSM3160813 Query DataSets for GSM3160813
Status Public on Dec 13, 2018
Title At_BT_rep2
Sample type SRA
 
Source name rosette leaves
Organism Arabidopsis thaliana
Characteristics ecotype: Col-0
tissue: leaves
developmental stage: 4 weeks post inoculation
infected with: BCTIV
Treatment protocol Plants were inoculated by agroinoculation, by delivery of agrobacteria carrying 1.4mer BCTIV-Siv clone (Soleimani et al., 2013) (accession no. JX082259) or pBIN19 as control (mock).
Growth protocol Plants were grown in soil in greenhouse at 20–28/16–20◦C (day/night), with a 16/8 h (light/dark) photoperiod and supplementary lighting
Extracted molecule genomic DNA
Extraction protocol Total nucleic acids were extracted from plant samples (100 mg) using the TLES method (Noris et al., 1996)
Before library preparation, genomic DNA was fragmented to an average insert size of 350bp using 18 cycles of 30 seconds each on a Bioruptor® Diagenode sonication device, split into three groups of 6 cycles with 2.5 minutes on ice between each set. The genomic DNA sequencing libraries were prepared using the TruSeq DNA PCR-Free LT Library Prep Kit following the manufacturer’s instructions (Illumina, San Diego, CA), starting from 1.1µg of sonicated DNA. Libraries were validated using the High Sensitivity D1000 screentape on the 2200 Tapestation instrument (Agilent technologies, Santa Clara, CA) and the LightCycler® 480 Instrument II using the LightCycler® 480 SYBR Green I Master mix (Roche, Basel, Switzerland).
 
Library strategy OTHER
Library source genomic
Library selection other
Instrument model Illumina NextSeq 500
 
Description Arabidopsis_BCTIV-infected_rep2
Data processing Library strategy: DNA-Seq
The raw reads were trimmed using Trimmomatic to remove adapter sequences. Reads with an averaged value of at least 15 in a 4 nt window were trimmed from both ends. After trimming, reads pairs with at least one mate shorter that 16 bp were discarded
The remaining sequences (in average 94% of raw reads) were aligned with segemehl v0.2.0 (Hoffmann et al., 2009), with 95% accuracy threshold, to the corresponding reference genome (A.thaliana TAIR10 and the B. vulgaris RefBeet-1.2.2) including also the sequence of the BCTIV genome (accession no. JX082259).
Split read junctions were called with testrealign.x (part of segemehl toolkit) with default parameters, and junctions with at least a coverage of 10 reads and comprising the virus genome were selected for downstream analysis using R (dplyr package)
Then, the reads spanning the 10 kb locus around each selected junctions were recovered together with their mates from the original fastq files, using samtools (Li et al., 2009) and BBMap (https://sourceforge.net/projects/bbmap/)
The filtered paired read lists were used as input in the A5_miseq pipeline (Coil et al., 2015) with default parameters, for reference-unbiased de novo assembly.
We then used blastn (BLAST v2.2.25+) with default parameters to filter only the assembled scaffolds matching both the reference and the BCTIV genome, to select minicircle candidates.
Genome_build: RefBeet-1.2.2
Genome_build: TAIR10
Genome_build: Beet curly top Iran virus-[Siv] (GenBank: JX082259)
Supplementary_files_format_and_content: Assembled scaffolds matching both the reference and the BCTIV genome were used to generate a list of sequences in fasta format. If none of the assembled scaffolds matched this alignment criteria, the file was not generated (this is the case of Arabidopsis samples and the B. vulgaris mock-inoculated plant).
 
Submission date May 28, 2018
Last update date Dec 13, 2018
Contact name Marco Catoni
Organization name University of Birmingham
Department School of Biosciences
Street address Edgbaston
City Birmingham
ZIP/Postal code B15 2TT
Country United Kingdom
 
Platform ID GPL19580
Series (1)
GSE114958 Virus-mediated export of chromosomal DNA in plants
Relations
BioSample SAMN09271198
SRA SRX4131434

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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