WGS Variant Calling: Variant Filtering and Annotation - Part 2 | Detailed NGS Analysis Workflow
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- čas přidán 29. 06. 2024
- This is the detailed workflow tutorial on how to filter variants and annotate them using GATK's Funcotator tool. In this video I discuss the need of filtering and annotating variants, the questions we may ask once we call variants, two approaches to filter variants i.e. Variant Quality Score Recalibration (VQSR) & hard filtering, the data sources available, genotype refinement and finally talking about Funcotator. Further, I demonstrate how to filter variants at the site level as well apply genotype filters and annotating variants using Funcotator. Lastly, I demonstrate how to wrangle the output from Funcotator and get the annotations in a tabular format which is helpful for further processing and visualization. I hope you find this video helpful! Leave your thoughts in the comment section below!
▸ Code:
github.com/kpatel427/CZcamsT...
▸ Part 1 of this video:
• WGS Variant Calling: V...
▸ VCF file format video:
• Understanding File For...
▸ GATK hard-filters recommendations for germline variants:
gatk.broadinstitute.org/hc/en...
Chapters:
0:00 Intro
0:43 Overview of variant calling steps
1:34 Questions we may want to ask after calling variants
3:00 Variant data sources
5:16 Two approaches to filter variants
8:38 Hard filtering: Site-level filtering
9:14 Hard filtering: Sample-level filtering
10:15 Genotype Refinement
10:54 Data used for today’s demo
11:22 Pre-requisites
12:22 Filtering SNPs
16:54 Filtering INDELS
17:35 Understanding output after filtering (FILTER column)
19:19 Select variants that passed filters
20:39 Exclude variants that failed genotype filters
25:49 GATK Funcotator tool
28:10 Funcotator data sources
29:59 Annotate variants using Functotator
32:07 Understanding output after annotation (Funcotator output)
33:45 Getting annotations into a tabular format
You can show your support and encouragement by buying me a coffee:
www.buymeacoffee.com/bioinfor...
To get in touch:
Website: bioinformagician.org/
Github: github.com/kpatel427
Email: khushbu_p@hotmail.com
#bioinformagician #bioinformatics #funcotator #variantcalling #variants #gatk #vcf #gvcf #haplotype #alleles #geneticvariants #mutations #gff3 #gff #gtf #sam #bam #phred #fasta #fastq #singlecell #10X #ensembl #biomart #annotationdbi #annotables #affymetrix #microarray #affy #ncbi #genomics #beginners #tutorial #howto #omics #research #biology #GEO #rnaseq #ngs
I usually never comment on anything, really, but I would be remiss if I didn't mention what a great job you are doing. There are not a lot of videos, or at least none I could really find, that make video tutorials on such specific topics like this. Really good job :) Thank you!
Thank you very much for Part 2 and especially for focusing on hard filtering as we requested you to do the video
Fantastic! Your presentation is clear and detail, helps me a lot in understanding the hard filtering process😊
i wish i found you sooner, i had to painstakingly browse though tones of GATK web pages to get me started with the Best practice guidelines. you made it clearer
Awesome tutorial! Thank you so much!
Thank you for this very informational video!
your tutorial really help me a lot to understand the workflow. If you have experience analyze gwas and also want to make the tutorial, I would appreciate it. Thank you so much ma'am ☺️
Thank you ma'am for sharing ur knowledge.... Happy to be ur student form online😊
You are great! Thanks for this amazing tutorial!
Thanks for the excellent tutorial👍
Great explanation!
you are great teacher.. thankyou.
thanks alot for this amazing video.
Thanks for ur amazing explanation. Plz keep posting more videos, can u make a video on how to use restapi, github, json, aws etc.
Fantastic work, thank you and it is always worth to watch your videos till the last second. Can one use this pipeline for WES as well?
Any script explanation for diplotype calling?, In the same way as you explained variant calling in a very clear way.
Good job
thank you very very very much
@22:00 Instead of doing all that, you can add '-select-genotype "GQ > 10 && DP > 10" \' to the SelectVariants commandline.
Thank you for sharing your knowledge with me. I understood everything well and was able to perform my analysis perfectly. I have a question regarding the view results. The command you provided was used only for SNPs. Should I use the same command for indels as well? The command (VIEW RESULTS STEP) is: "cat ${results}/analysis-ready-indels-filteredGT-functotated.vcf | grep " Funcotation fields are: " | sed 's/|/\t/g' > ${results}/output_curated_variants_indels.txt" and "cat ${results}/output_indels.table | cut -f 5 | sed 's/|/\t/g' >> ${results}/output_curated_variants_indels.txt". Please let me know. Thank you once again for your help.
This is a great tutorial !.
Can you please make a video on "Call somatic mutations using GATK4 Mutect2"?? and a video on "maftools" - R package to analyze somatic .vcf files??"
Highly appreciated !
I will surely consider covering these topics. Thank you for the suggestion:)
Thank you that was helpful, but what if I have the WES data in form of Excel sheet how can I filter my variants? and is there a tool or a program can do that?
Thanks very much for the video. Could you continue to explain Call somatic mutations with Mutect2?
I will surely consider making a video on mutect2
@@Bioinformagicianyes ... requesting a tutorial for somatic variant calling workflow....thank you
@Bioinformagician please.....As soon as possible.
Could you please make a video for GTAK-mutect2 variant calling?
Does this include somatic and germline variant calling workflow?
how to set those filters ? what do all do we have to consider ?
Mam, please upload the visualization part
Why don't you just merge indels and snps in one main file and annotate this one main file right away?
You are amazing. Thanks!!!