Introduction

Genome analysis of Botryotinia fuckeliana (anamorph Botrytis cinerea) strain BcDW1.

Dataset Description

Botrytis cinerea (teleomorph Botryotinia fuckeliana) is an opportunistic pathogen with a broad host range and is particularly aggressive on fleshy fruit, such as tomato, strawberries, and grape berries. DNA from an axenic BcDW1 culture was extracted using a modified cetyltrimethylammonium bromide (CTAB) method, and 6.9 Gb of Illumina sequence reads was generated to achieve >100× coverage of the genome.

• Organism: Botryotinia fuckeliana (Botrytis cinerea BcDW1).

• Instrument:  Illumina HiSeq 2000.

• Layout: Paired-end.

Publication

Blanco-Ulate B, Allen G, Powell AL, Cantu D. Draft Genome Sequence of Botrytis cinerea BcDW1, Inoculum for Noble Rot of Grape Berries. Genome Announc. 2013 May 23;1(3):e00252-13. doi: 10.1128/genomeA.00252-13. PMID: 23704180; PMCID: PMC3662820.

Original Data

• NCBI BioProject: PRJNA188482.

• SRA Experiment: SRR680162.

• NCBI Assembly: GCA_000349525.1.

Bioinformatic Analysis

1- DNA-Seq de novo Assembly

Application

DNA-Seq de novo Assembly (ABySS).

Input

• Illumina sequencing data in FASTQ format (SRR680162_1.fastq.gz and SRR680162_2.fastq.gz).

Parameters

• Input Reads: [Paired-End] SRR680162_1.fastq.gz and SRR680162_2.fastq.gz

• Upstream Files Pattern: _1

• Downstream Files Pattern: _2

• Use Additional Data: false

• K-mer Size: 64

• Use Paired de Bruijn graph: false

• Minimum Alignment Length: 40

• Hash Functions: 1

• K-mer Count Threshold: 2

• Unitigs Fasta: unitigs.fasta

• Contigs Fasta: contigs.fasta

• Scaffolds Fasta: scaffolds.fasta

• Save Graph Files: true

Execution Time

70-90 minutes.

Output

• unitigs.fasta: FASTA file containing assembled unitig sequences.

• contigs.fasta: FASTA file containing assembled contig sequences.

• scaffolds.fasta: FASTA file containing assembled scaffold sequences.

• report_abyss.box: Report about DNA-Seq de novo Assembly results.

• nx_plot.box: Line chart about DNA-Seq de novo Assembly results.

• assembly-contigs.dot: Text file containing the contig overlap graphs in the GraphViz DOT syntax.

• assembly-scaffolds.dot: Text file containing the scaffold overlap graphs in the GraphViz DOT syntax.

2- Repeat Masking

Application

Repeat Masking.

Input

• Assembled genomic scaffolds in FASTA format (from the 1- DNA-Seq de novo Assembly step).

Parameters

• Input Sequences: scaffolds.fasta

• Search Engine: RMBlast

• Repeat Database: Dfam Consensus

• Species: 40559 Botrytis cinerea

• Speed/Sensitivity: Default

• Masking Options: Soft masking (lowercase)

• Apply Divergence Cutoff: false

• Only Alu elements: false

• Type of repeat: interspersed,simple_low

• Not mask RNA genes: false

• Output FASTA: masked_sequences.fasta

Execution Time

10-15 minutes.

Output

• masked_sequences.fasta: FASTA file containing repeat masked sequences.

• scaffolds_gff.box: GFF project containing repeat coordinates.

• report_repeat_masking.box: Report about Repeat Masking results.

• repeat_type_distribution.box: Pie chart showing the abundance of each type of detected repeats.

3- Gene Finding

Application

Eukaryotic Gene Finding by AUGUSTUS.

Input

• Repeat masked sequences in FASTA format (from the 2- Repeat Masking step).

Parameters

• Input Sequences: masked_sequences.fasta

• Closest Species: Botrytis cinerea [Fungi - Ascomycota - Leotiomycetes]

• Strand: Both Strands

• Allowed Gene Structure: Partial

• Output Genomic Features: introns,start,stop

• Ignore Strand Conflicts: false

• UTR Prediction: false

• No In-frame Stop Codons: false

• Stop Codons Excluded From CDS: false

• Softmasked Sequences: true

• Sample: 100

• Alternatives From Sampling: false

• Gene Finding Mode: Ab Initio Prediction

Execution Time

5-10 minutes

Output

• cds.box: Sequence project containing predicted gene sequences.

• protein.box: Sequence project containing predicted protein sequences.

• coordinates.box: GFF project containing predicted gene coordinates.

• report_eukaryotic_gene_finding.box: Report about Eukaryotic Gene Finding results.

• cds_length_distribution.box: Bar chart showing the length distribution of the predicted genes.

4- BLAST & InterProScan

Application

CloudBLAST & InterProScan Annotation.

Input

• Predicted gene sequences in an OmicsBox project (from the 3- Gene Finding step).

Parameters

Execution Time

~ 1 hour.

Output

• blast_ips_protein.box: Project containing predicted protein sequences along with Blast and IPS results.

Workflow