Building and validating a variant calling pipeline

1. From NGS to INDELs
Building and Valida+ng a Variant
Calling Pipeline for NGS Data
Jatinder Singh

2. Overview of Next Genera0on
Sequencing
2
DNA Sample
NGS Instruments
- Sequencing
- Base Calling
NGS Data Analysis
- Alignment
- Variant Calling: SNPs, INS, DEL, INV, DUP
read1: ATCGCTTC
read2: TATCGCTT
read3: CTTCCATG
read4: CGCTTCCA
read5: CCATGACG
read6: TTCCATGA

3. Insertions and
Deletions (INDELs)
3
GC GA
T A
GC GA
GC GA
GTC A A G
GC GA C GG
Insertion
Deletion

4. 4
INDEL Calling Programs
- HaplotypeCaller
- Freebayes
- Pindel
- Lumpy
▫ Simulate NGS dataset for validation using VarSim
▪ Illumina PE data, 10x, 20x and 40x coverage
▪ 10,000 small INDELs
▪ 2000 large INDELs
Validation

5. 5
Program Performance at
10x Coverage

6. 6
Program Performance at
40x Coverage

7. INDEL Calling Pipeline
7
NGS Data
(FASTQ files)
INDEL Calling
Alignment
(BWA-mem)
Low Coverage
Sequence Data
High Coverage
Sequence Data
- HaplotypeCaller
- Lumpy
- Pindel

8. About me
8
Questions?
Jatinder Singh
jatinder.kang@gmail.com