Nature Genetics Vol.36 Sept 2004

Detection ofLarge-scale VariationIn the Human GenomeIafrate, Feuk, Rivera, Listewnik, Donahoe, Qi, Scherer, Lee

anyimagehere

Title Page

Trevor PughMEDG505 – Genomic AnalysisUniversity of BC

Large-scale Copy-Number Variations: 5 W’s & How

What are Large-scale Copy Number Variations?

Who have Large-scale Copy Number Variations?

Where are Large-scale Copy Number Variations?

Why are there LCVs?

When did LCVs arise?

How are LCVs generated?

What are Large-scale Copy-Number Vars.?

@ gains or losses of kb’s-Mb’s of genomic DNA

@ seen in phenotypically normal individuals

> may not be direct cause of genetic disease

> may give rise to disease?chromosomal rearrangementssubtle phenotypic variation (gene expression)

@ NOT limited to intergenic or intronic regions

Who have Large-scale Copy-Number Vars?

1 1 1 1 1 1 1 1 1 1 2 1 1 1

Deletion!Five steps of CGH@ Isolation of Genomic DNA from two patients/sources

@ Digestion of Genomic DNAinto short fragments

@ Labeling of each sample withDIFFERENT detectable dyes

@ SIMULTANEOUS Hybridizationof labeled probes to targets

@ Quantification of amount ofhybridization to each target

@ applied array CGH to 55 unrelated individuals

@ spots are large DNA fragments ‘tiled’ ~1MB across the genome low resolution, captures 12% of genome

@ applied array CGH to 55 unrelated individuals> 39 healthy phenotypes (normal karyotype)> 16 with known chromosomal abnormalities... against a pool of (different) normal controls

@ detected *all* known LCVs in 16 abnormal samples> good sensitivity and specificity

@ on average, detected 12.4 LCVs in normal controls!> mostly single large insert clones

up to 2Mb affected!

Who have Large-scale Copy-Number Vars?

@ on average, detected 12.4 LCVs in normal controls

(78)

(24)

153 >1 individual

>10% of individuals

Low frequency

# of LCVs found in:

(False Positive rate <1 per 5,264 clones)

@ 26 mapped to previously recognized segmental dupes@ 13 reside within 100kb gaps of current human genome sequence presentation

complicates assembly of final HGP sequence

102

Who have Large-scale Copy-Number Vars?

Everywhere!

RED = copy gainsBLUE = copy losses

Where are Large-scale Copy-Number Vars?

@ across the genome involving as much as 2MB

@ not limited to intergenic or intronic regions> can include duplicate genes!

@ confined to localized chromosomal regions

Where are Large-scale Copy-Number Vars.?

OR

> NOT duplication events of other chromosomes> tandem copy-number changes

@ tandem copy-number changes> visualized using FISH

Where are Large-scale Copy-Number Vars.?

RELATIVE: Losses Normal Gains

signals only from sister

chromatids

> CN gains show more head-to-tail gene copies

RED =5’ headGREEN =3’ tail

& fibre-FISH

6 duplicates 9 duplicates 12 duplicates

Where are Large-scale Copy-Number Vars.?@ tandem copy-number changes

> verified by QT-PCR

> ratios track FISH & CGH ratios almost perfectly!

Why do Large-scale Copy-Number Variations Exist?

@ Unknown!

When did Large-scale Copy-Number Variations Arise?

“Large-Scale Variation Among Human and Great Ape Genomes Determined by Array Comparative Genomic Hybridization”Locke, Segraves, Carbone, Archidiacono, Albertson, Pinkel, EichlerGenome Research Vol 13, Issue 3, 347-357, March 2003

@ gene-rich regions susceptible to CN changes between humans and great apes

@ local repatterning of hominoid chromosomes in euchromatic regions

> duplication-driven mechanism of genome evolution

@ Unknown!

How do LCVs Impact Us?

@ new explanations for individual uniqueness> lineage studies?> forensics?

@ increased gene copy number associated with increased susceptibility to diseases?

> use as genetic markers?

@ added level of complexity to the genome> comparative genomics?> phylogenetics?

@ simplification of the genome> less regulatory networks, more ‘dosage effects’

Summary:Large-scale Copy-Number Variations (LCVs)

What?@ gains or losses of kb’s-Mb’s of tandem genomic sequences

Who?@ varies from person to person, 12.4 LCVs on average

Where?@ throughout the genome, localized to chromosomal regions@ not limited to intergenic or intronic regions

Why?@ Unknown! Conjecture: Gene evolution? Regulatory?

When?@ Unknown! Conjecture: Complex organisms? Plant/Animal?

How?@ new basis for human uniqueness & treatment@ new ways of interpreting the genome and its interactions

Questions?

Questions@ Why do LCVs exist? When did they arise?

@ Are these LCVs random? If not, what types of genes would have copy number variation?

@ What possible mechanisms could create LCVs?

@ Is this a genomic simplification or an added level of complexity?

@ What are the implications of LCVs for current technologies, databases, and assemblies?

> PCR, gene expression, genetics?> GenBank, OMIM?> Human Genome Project?

Extra Figures