Accurate variant calling in NGS data is a critical step upon which virtually all downstream analysis and interpretation processes rely. Germline analysis in family trios and somatic analysis of tumor-normal pairs.
Variant calling is a crucial step in the analysis of genomic data, particularly in the field of genetics and genomics. It involves identifying and cataloging differences (variants) in the genetic material (DNA or RNA) when comparing it to a reference genome or another sample. These variants can include single nucleotide changes (SNPs), insertions, deletions, and structural variations.
Variant calling has become the basis of downstream analysis for all the genetically predisposed diseases, like diabetes, downs syndrome, autism, arthritis and to much extent cancer.
Variant analysis steps include a robust NGS alignment pipeline and pre-processing pipeline, variant calling, filtering the variants and manual review.
Variants such as deletions or insertions can be called by any given package but structural and copy number variants should be analyzed by multiple softwares.
We at genomebeans thrive to provide you with robust and reliable analysis on which you can carry forward your downstream analysis.
Raw data is generated from laboratory is used as sample for sequencing data analysis.
The raw data is converted into .fastq format for its quality check and is tested on different parameters to identify sequence purity.
On the basis of sequence quality it is trimmed and adapter or any mismatch pair are removed.
Index is prepared with reference file that holds information about gene structure.
The sample is mapped to reference based on the index generated, to identify the reads mapped to a given gene in the reference.
Unsorted aligned BAM sample files thus generated are further sorted for gene counts.
We use the GATK HaplotypeCaller to perform variant calling. The HaplotypeCaller is capable of calling SNPs and indels simultaneously via local de-novo assembly of haplotypes in an active region.
This step separates SNPs and Indels so they can be processed and analyzed independently. Various hard filters are applied to further filter the variants for he highest quality and prep for annotation.
Used funcotator tool for GATK that uses pre-packaged vcf database from GRCh38 and hg19. Depending on the need we can use SNPeff, gnomad, etc to annotate the variants.
We use IGV for visualization.
