1 Commentary 1.Do not get too worried about "methods" and details. I fully expect there to be concepts and techniques that you simply are not going to know. 2.Try to figure out the "big picture." 3.Finding papers (from campus you will have access to a lot more journals) PubMed is better for finding more recent papers (ordered by date). Google/scholar is nice for finding papers that reference a paper When capturing figures from papers, blow up the figure as large as possible, capture, then shrink to fit slides (not the other way around). Spelling, grammar, syntax -- all count.

2 Segmental Duplications and Copy-Number Variation in the Human Genome Sharp et al. Am. J. Hum. Genet. 77:78-88, 2005 presented by: Terry Braun

3 Abstract Human genome contains blocks of duplicated sequence (segmental duplications) Identified 130 potential rearrangement hotspots (bioinformatically?) Created a "microarray" to assess copy-number variations Examined 47 "normal" individuals Identified 119 regions of copy-number polymorphism (CNP)

4 Outline Background –CGH –Segmental duplications Materials and Methods –very brief Results –CNPs in the population –Validation Discussion

5 Background Segmental duplications –blocks of DNA -- 1kb to 400 kb 5% of genome is duplicated Implicated in more than 25 genomic disorders Array comparative genomic hybridization (CGH)

6 Background (cont) Referenced -- Representational Oligonucleotide Microarray Analysis (ROMA) Nice description on wikipedia Main difference -- uses restriction enzymes on the test and reference genome -- which improves efficiency

7 Background (cont) Previous work had limited coverage of whole genome. Hypothesis: regions flanked by segmental duplications are susceptible to rearrangement -- "hotspots" of genome instability that results in "copy- number variations"

8 Background (cont) Utilized knowledge from previous studies on genomic disorders and copy number variations: –regions flanked by duplications –duplications > 10K –>95% sequence identity –region spanned is 50kb to 10Mb Searched genome for regions with these characteristics (in silico) Disappointed in lack of details

9 Figure 1

10 Material and Methods 130 regions found Selected "BACs" –Bacterial artificial chromosomes –Technique for keeping and duplicating portions of the genome –Used by human genome project –100kpb kbp Analysis used to select BACs

11 BAC Selection Priority 1.BACs contained in "hotspot" 2.BACs overlapping segmental duplications 3.flanking BACs in peripheral unique sequence as local control Selected 2,194 BACs (192 single copy, randomly selected)

12 Microarry Comparative Genome Hybridization glass slide (or silicon waffer) DNA is spotted (or grown) High density (1000's to millions of "spots" or probes) Millions of copies per "spot"

13 Source: graphics by Jiang Long

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15 Results Validation –true positive strong correlation to BAC's that "lit up" in 3 individuals with known CNV's, and proportional to number of copies –false positive 4 self-vs-self (assuming they used the BACs???) threshold of log2 hyb ratios deviating > 2 standard deviations from mean correspond to FPR ~3 per 4000

16 Results 160 variant BACs in 47 individuals (corresponding to 119 nonredundant regions)

17 Copy Number Polymorphisms (CNP)

18

19 More Validation -- Individual

20 FISH Validation – Segmental Duplication BAC

21 Validation – Chrm 8 BACs

22 Analysis of CNPs 15.7% of "hotspot" regions were variant 4.1% non-hotspot regions were variant => 3.8-fold enrichment Chi^2 = 73.8 P < CNV regions were "not particularly gene poor."

23 Discussion "4-5-fold enrichment of CNPs flanked by SD's Supports notion that regions are predisposed to rearrangement Multi-ethnicity speculations –evolutionarily ancient, prior to separation of groups, OR –recurrent events that occurred independently in multiple founders

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25 Supplemental Data

26 Conclusions Duplications may be impediment to genome assembly UCSC genome view of CNP neticsdiv/anomaly%20register/ Hotspots represent excellent candidate sites of recurrent rearrangements associated with novel genomic disorders

27 Summary Comparative genome hybridization 47 individuals Copy number polymorphisms found Enriched for CNPs by examining segmental duplications (a bioinformatics approach) Appears to be convincing Resolution leaves a lot to be desired Difference between “hotspot” and “segmental duplications” regions not clear