Jan2016 bina giab

1. Validating, Enhancing and Using
GiaB Reference Materials
1
GiaB Workshop, Stanford University
January 29, 2016
Bina Technologies, Roche Sequencing
Marghoob Mohiyuddin
Mohammad Sahraeian
Hugo Y. K. Lam

2. Background
2

3. For Research Use Only. Not for use in diagnostic procedures.
What we do
3

4. For Research Use Only. Not for use in diagnostic procedures.
Collaborative scientific innovation
4
SeqAlto
VarSim

5. Benchmarking variant-calling

6. For Research Use Only. Not for use in diagnostic procedures.
The VarSim framework

7. For Research Use Only. Not for use in diagnostic procedures.
The VarSim framework
Simulate and validate whole-
genomes
Both somatic and germline
simulation supported
Comprehensive simulation and
validation of multiple kinds of
variants
SNPs, small Indels, SVs

8. For Research Use Only. Not for use in diagnostic procedures.
Assessing variant-calling
VarSim simulation of 50x Illumina 2x100bp reads for NA12878
Error profiles from NA12878 platinum genomes sample
SNVs and small indels from GiaB high-confidence set
SVs from multiple sources
Deletions from 1000Genomes
Insertions randomly sampled from DGV and sequences from HuRef insertion
sequences
Inversions, duplications randomly sampled from DGV
8

9. For Research Use Only. Not for use in diagnostic procedures.
Small variant calling accuracy

10. Validating GiaB gold set

11. For Research Use Only. Not for use in diagnostic procedures.
GiaB SVs
11
GiaB released high-confidence SVs for NA12878
2676 deletions, 68 insertions
Trio sequences available from Illumina Platinum Genomes
MetaSV calls SVs by integrating from multiple methods
http://bioinform.github.io/metasv/
Validation of GiaB gold set using MetaSV trio analysis ensures
quality of GiaB gold set

12. For Research Use Only. Not for use in diagnostic procedures.
MetaSV trio analysis
12
Validate by analyzing trios (50x coverage)
MetaSV calls for parents (NA12891, NA12892)
MetaSV calls for NA12878
Criteria
Deletions >= 100bp considered (2,348/88% in GiaB)
Reciprocal overlap of 50%
GiaB deletion validated if
Detected by MetaSV in any parent (multiple samples)
Detected by MetaSV as high-confidence in NA12878 (multiple methods)
Reported in previous literatures

13. For Research Use Only. Not for use in diagnostic procedures.
MetaSV: an ensemble approach
13

14. For Research Use Only. Not for use in diagnostic procedures.
MetaSV workflow
14
Merge SVs from multiple
methods
Multiple methods  high-conf.
8 SV callers supported
Enhanced insertion detection
Soft-clip analysis + assembly
Assembly and alignment to
refine breakpoints
Supports Del, Ins, Inv, Dup,
Trans.
Supported in bcbio

15. For Research Use Only. Not for use in diagnostic procedures.
GiaB deletion validation
15
Total
Validated
Total not
validated
Additionally
Validated
0
(0%)
2,348
(100%)
0
(0%)
2,302
(98.0%)
46
(2.0%)
2,302
(98.0%)
2,306
(98.2%)
42
(1.8%)
4
(0.2%)
2,342
(99.7%)
6
(0.3%)
36
(1.5%)
GiaB HC
GiaB HC Validated by Parents (MetaSV ALL)
GiaB HC Validated by Child (MetaSV PASS)
GiaB HC Validated by Child (curated)

16. For Research Use Only. Not for use in diagnostic procedures.
Mendelian validation
16
● MetaSV High Quality Trio Deletions:
○ Mendelian Inheritance Consistency with Genotypes
○ Pass in Child and ALL in Parents
○ Considering no call as reference call
MetaSV Trio Dels
2,582
GiaB HC Dels
2,348
GiaB
Private
222
Common
2,126
MetaSV Private
456
(142 not in
literature)
MetaSV PASS Dels
2,671
96.7% are Mendelian
consistent
(98.7% if ignoring
genotypes)

17. For Research Use Only. Not for use in diagnostic procedures.
Trio analysis summary
17
GiaB SVs have a high validation rate using MetaSV trio analysis
Only 6 unvalidated SVs do not have strong support in IGV or SVVIZ
 GiaB deletions of high quality
Almost all (up to 98.7%) MetaSV PASS calls are Mendelian
consistent making them high-quality
Significant number (456) of MetaSV trio calls not in GiaB
Possibly missed due to stringent GiaB requirements since 321 of those in
literature
MetaSV trio validation can help validate and extend the gold set

18. Enhancing GiaB gold set

19. For Research Use Only. Not for use in diagnostic procedures.
Work on Jewish trio
19
Enhanced MetaSV
Assembly to get evidence for and refine all kinds of SVs
Optimizations to speed up analysis
Calls for Jewish trio submitted
Will help build high-confidence SV calls for other trios

20. For Research Use Only. Not for use in diagnostic procedures.
MetaSV and Parliament
20
2240(MetaSV,79.7%)
4490(Parliament,82.1%)
569(20.3%)
977(17.9%)
MetaSV total
2809
Parliament
total 5467
(after uniq)
Reciprocal overlap of 50% was used, no genotype matching performed. With 90%
reciprocal overlap, 75.9% MetaSV calls and 54.0% Parliament calls overlapped.

21. For Research Use Only. Not for use in diagnostic procedures.
Summary
21
GiaB high-confidence calls regularly used to assess variant-calling
pipelines
For both simulated and real data
MetaSV was used to perform validation for GiaB deletion SVs for
NA12878
Trio analysis can also be applied to the Jewish and Han samples
Contributing MetaSV calls for Jewish trio to help build better SV
gold sets

22. For Research Use Only. Not for use in diagnostic procedures.
Acknowledgements
22
Bina: Mohammad Sahraeian, Jian Li, John Mu
GiaB: Justin Zook, Hemang Parikh