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training:dudijon:galaxy [2021/12/14 18:12] slegras [9.2 Run the workflow DNA-seq data analysis] |
training:dudijon:galaxy [2022/12/09 15:37] (current) slegras |
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| ^ Instructor ^ Stephanie Le Gras ^ | ^ Instructor ^ Stephanie Le Gras ^ | ||
| ^ Duration | 3.5 hours | | ^ Duration | 3.5 hours | | ||
| - | ^ Content | {{:training:dudijon:introgalaxy_2020_compressed.pdf|Description of the key features of Galaxy (Lecture)}} | | + | ^ Content | {{:training:dudijon:introgalaxy_2021_compressed.pdf|Description of the key features of Galaxy (Lecture)}} | |
| ^ ::: | Practical session on basic features of Galaxy (Hands-on) | | ^ ::: | Practical session on basic features of Galaxy (Hands-on) | | ||
| ^ Prerequisites | None | | ^ Prerequisites | None | | ||
| Line 27: | Line 27: | ||
| ++++ | ++++ | ||
| - | ==== - Import data into Galaxy ==== | + | ==== - Import files from your computer to Galaxy ==== |
| - | === - Import files from your computer to Galaxy === | + | |
| - | - Download the two files **CRN-107_11-R1.fastq.gz** and **CRN-107_11-R2.fastq.gz** following this [[https://seafile.igbmc.fr/d/345d7581237d4295bf2c/|link]]. | + | |
| - | - Import them to your history called “DNA-seq data analysis” | + | |
| - | * The genome is: Human (hg19) | + | - Download the file “**sample.bed.gz**” following this [[https://seafile.igbmc.fr/d/1adaad8f80394182a784/|link]] and upload it to Galaxy. |
| - | * The format: <auto detect> | + | |
| - | + | ||
| - | ++++ Answer | | + | |
| - | - Click on "Upload Data" | + | |
| - | - Drag and drop the two fastq files “**CRN-107_11-R1.fastq**” and "**CRN-107_11-R2.fastq**" | + | |
| - | - Select/Enter Genome for both datasets as: hg19 | + | |
| - | - Click on Start | + | |
| - | + | ||
| - | {{:training:dudijon:03.1-LaunchDragAndDrop.png?|}} | + | |
| - | {{:training:dudijon:03.2-UploadFastqFiles.png?|}} | + | |
| - | ++++ | + | |
| - | + | ||
| - | ==== - Remove a dataset ==== | + | |
| - | === - Import a file from your computer === | + | |
| - | - Download the file “**sample.bed.gz**” following this [[https://seafile.igbmc.fr/d/345d7581237d4295bf2c/|link]] and upload it to Galaxy. | + | |
| * The genome is: Mouse (mm9) | * The genome is: Mouse (mm9) | ||
| * The format is: bed | * The format is: bed | ||
| Line 62: | Line 44: | ||
| ++++ | ++++ | ||
| + | ==== - Remove a dataset ==== | ||
| - Remove the dataset **sample.bed** from your history by clicking on the button | - Remove the dataset **sample.bed** from your history by clicking on the button | ||
| - You are told that your history is empty. Look at the size of your history | - You are told that your history is empty. Look at the size of your history | ||
| - | - Click on “**deleted**” in the top of the history panel (below the history name). Remove definitely the file from the disk by clicking on "**Permanently remove it from disk**”. | + | - Click on “**deleted**” in the top of the history panel (below the history name). Remove definitely the file from the disk by clicking on "**Supprimer définitivement du disque**”. |
| - Click on “hide deleted” | - Click on “hide deleted” | ||
| ==== - Running a tool ==== | ==== - Running a tool ==== | ||
| + | - Download the two files **CRN-107_11-R1.fastq.gz** and **CRN-107_11-R2.fastq.gz** following this [[https://seafile.igbmc.fr/d/1adaad8f80394182a784/|link]]. | ||
| + | - Import them to your history called “DNA-seq data analysis” | ||
| + | * The genome is: Human (hg19) | ||
| + | * The format: <auto detect> | ||
| + | |||
| + | ++++ Answer | | ||
| + | - Click on "Upload Data" | ||
| + | - Drag and drop the two fastq files “**CRN-107_11-R1.fastq.gz**” and "**CRN-107_11-R2.fastq.gz**" | ||
| + | - Select/Enter Genome for both datasets as: hg19 | ||
| + | - Click on Start | ||
| + | |||
| + | {{:training:dudijon:03.1-LaunchDragAndDrop.png?|}} | ||
| + | {{:training:dudijon:03.2-UploadFastqFiles.png?|}} | ||
| + | ++++ | ||
| + | |||
| - Use the tool “FastQC Read Quality reports” to compute quality analysis on the datasets “**CRN-107_11-R1.fastq**” and "**CRN-107_11-R2.fastq**" | - Use the tool “FastQC Read Quality reports” to compute quality analysis on the datasets “**CRN-107_11-R1.fastq**” and "**CRN-107_11-R2.fastq**" | ||
| - Use default parameters. | - Use default parameters. | ||
| Line 95: | Line 93: | ||
| * CRN-107_11-R1.fastq | * CRN-107_11-R1.fastq | ||
| * CRN-107_11-R2.fastq | * CRN-107_11-R2.fastq | ||
| - | * CaptureDesign_chr4.bed (download it from [[https://seafile.igbmc.fr/d/345d7581237d4295bf2c/|here]]) | + | * CaptureDesign_chr4.bed (download it from [[https://seafile.igbmc.fr/d/1adaad8f80394182a784/|here]]) |
| Import missing files from the data library "**DNA-seq test datasets**" | Import missing files from the data library "**DNA-seq test datasets**" | ||
| Line 123: | Line 121: | ||
| - Limit analysis to regions in this BED dataset: CaptureDesign_chr4.bed | - Limit analysis to regions in this BED dataset: CaptureDesign_chr4.bed | ||
| - __SnpEff__ Variant effect and annotation | - __SnpEff__ Variant effect and annotation | ||
| - | - Sequence changes (SNPs, MNPs, InDels): **output of GATK Haplotype Caller (VCF)** | + | - Sequence changes (SNPs, MNPs, InDels): **output of FreeBayes (VCF)** |
| - Input format: VCF | - Input format: VCF | ||
| - Output format: VCF (only if input is VCF) | - Output format: VCF (only if input is VCF) | ||
| Line 143: | Line 141: | ||
| === - Rename the workflow "DNA-seq data analysis" === | === - Rename the workflow "DNA-seq data analysis" === | ||
| ++++ Answer | | ++++ Answer | | ||
| - | {{:training:dudijon:04-manageworkflow.png?|}} | + | {{:training:dudijon:05-editorrunworklow.png?|}} |
| - | Now your can edit or run the workflow: | ||
| - | |||
| - | {{:training:dudijon:05-editorrunworklow.png?|}} | ||
| ++++ | ++++ | ||
| Line 174: | Line 169: | ||
| - __Samtools flagstat__ to compute mapping statistics (after BWA mem) | - __Samtools flagstat__ to compute mapping statistics (after BWA mem) | ||
| - | - __Filter__ to select aligned reads with a mapping quality >= 20 (after MarkDuplicates) | + | - __Filter SAM or BAM, output SAM or BAM__ to select aligned reads with a mapping quality >= 20 (after MarkDuplicates) |
| - __Samtools flagstat__ to compute mapping statistics after removing reads with low mapping qualities (after Filter) | - __Samtools flagstat__ to compute mapping statistics after removing reads with low mapping qualities (after Filter) | ||
| Line 180: | Line 175: | ||
| - __Flagstat__ tabulate descriptive stats for BAM dataset | - __Flagstat__ tabulate descriptive stats for BAM dataset | ||
| - BAM File to Convert: **output of BWA mem** | - BAM File to Convert: **output of BWA mem** | ||
| - | - __Filter__ BAM datasets on a variety of attributes | + | - __Filter SAM or BAM, output SAM or BAM__ files on FLAG MAPQ RG LN or by region |
| - | - BAM dataset(s) to filter: **output of Picard MarkDuplicates** | + | - SAM or BAM file to filter: **output of Picard MarkDuplicates** |
| - | - Select BAM property to filter on: mapQuality | + | - Minimum MAPQ quality score: **20** |
| - | - Filter on read mapping quality (phred scale): **>=20** (this exact expression, including ">="!) | + | |
| - __Flagstat__ tabulate descriptive stats for BAM dataset | - __Flagstat__ tabulate descriptive stats for BAM dataset | ||
| - BAM File to Convert: **output of Filter** | - BAM File to Convert: **output of Filter** | ||
| Line 215: | Line 209: | ||
| ++++ Answer | | ++++ Answer | | ||
| - | 561598 - 531417 = 30181 | + | 561598 - 530355 = 31243 |
| ++++ | ++++ | ||
