sept2016 sv bio_nano

PowerPoint Presentation

Genome in a Bottle UpdateSeptember 15th, 2016

#2015 BioNano Genomics

Algorithms Convert Images into MoleculesAssembly Algorithms Align Molecules de novo for Constructing Consensus Genome MapsCross-Mapping Across Multiple Samples or to a ReferenceHigh Throughput, High Resolution Imaging Gives Contiguous Molecules up to Mb LengthAutomated SV DetectionGap SizingGenome FinishingIsolate High Molecular Weight DNALabel Specific Sequences Across the Entire GenomeTransfer Labeled DNA into Cartridge for Scanning

Insertion

Irys Workflow- Overview

Load, Linearize & Image Labeled DNA in Repeated Cycling to Scan Whole Genome


Irys Applications

Structural Variation (SV) discoveryCompare to a reference or gold standard, looking for changes in the patterns:Shifts in barcode patterns reveal insertion (addition), deletion (subtraction), inversion (re-orientation), translocation of genome segments.Hybrid ScaffoldingBioNano maps are used as a scaffold:Sequencing contigs are converted in silico into molecular barcodes by highlighting the same sequence motifsThese sequencing based barcodes are then aligned to the BioNano maps

Two Fundamental Applications

#2015 BioNano GenomicsRead the slide 3

Label Site Gain/ LossBalanced SVsTandem ArrayDeletionInsertion

Cross-Map Analysis: Look for Similarities and Differences Between Maps

Deletion

Insertion

26.6kb 21.0kb

IrysView SV Detection

BCRABL1 Chr 9Chr 22

~200kb~70kb


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Simulated Diploid Genome Sensitivity and PPV EstimateBioNano SV calls in two homozygous hyditidaform mole cell lines were used as a gold standardVariants that are the same in both lines are considered homozygous and variants found in only one line are considered heterozygous in the mixtureWe do not have a measurement of FN rate related to missing coverage (e.g. at fragile sites) in this set up


BioNano sizePacBio size (bp)Size Concordance for Insertions and Deletion Between PacBio and BioNano SVs

BioNano sizePacBio size (bp)Pac Bio calls from: Chaisson et al., Resolving the complexity of the human genome using single-molecule sequencingDeletionsInsertionsGood size concordance between BioNano and PacBio SVs facilitates cross validation of findings.


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New SV Calls Better Heterozygous SV Detection, Confidence ScoresSampleNickaseInput dataDiploid assembly sizeGenome map N50InsertionDeletionFatherNt.BspQI319 Gb5.24 Gb4.46 Mb44361935MotherNt.BspQI385 Gb5.22 Gb3.93 Mb50472210SonNt.BspQI508 Gb5.52 Gb2.55 Mb36261775

Check son-specific SVs:low assembly, alignment (repetitive, ambiguous), variant scoresFP in childFN in parentsputative de novo SVsAggressive SV calling

231132291034 son-specific SVs5238036793089FatherMother


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FP variant in the son low local coverage

1.055 MbpinsHeterozygous Insertion on chr18:31,368,891-31,419,488 size=159.8 kbp

Note that the coverage at the variant region is abnormally low. Hence this is a false call, and is filtered by automated molecule alignment checkSon mapSon mapchr18Son molecule pileup

#2015 BioNano GenomicsFN in a parents assembly (1)

Son302 kbpchr8

MotherFather

A heterozygous deletion found only in the son by assembly comparison. chr8:5,594,272-5,609,910size: 10.2 kbpOn the mothers assembly (blue arrow), there seem to be some kind of aberration, but it does not match the sons deletionDel

#2015 BioNano GenomicsFN in a parents assembly (2)Mothers molecule aligned to the sons genome map

SonMothermolecules302 kbpA heterozygous deletion found only in the son by assembly comparison. chr8:5,594,272-5,609,910size: 10.2 kbp

Note that the mothers molecules actually align well to the sons genome map


231132293 son-specific SVs5238036793089FatherMotherSV shared among trio members792 with low assembly, alignment (repetitive, ambiguous), variant scores20 FP in child219 FN in parents3 putative de novo SVs

SampleNickaseInput dataDiploid assembly sizeGenome map N50InsertionDeletionFatherNt.BspQI319 Gb5.24 Gb4.46 Mb44361935MotherNt.BspQI385 Gb5.22 Gb3.93 Mb50472210SonNt.BspQI508 Gb5.52 Gb2.55 Mb36261775


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SummaryFully de novo genome map assembly for genome structureValidation of sequence assembly by orthogonal verification Hybrid scaffolding of sequence assembliesStructural variation detectionStructural variation validation by genome map and single molecule alignment

#2015 BioNano GenomicsAcknowledgements

Genome in a bottle consortiumBioNano GenomicsAndy PangJoyce LeeErnest LamTiff LiangThomas AnantharamanKhoa PhamWill StedmanHan CaoMt. SinaiAli Bashir


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sept2016 sv bio_nano

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