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| Status |
Public on Feb 01, 2010 |
| Title |
Highly efficient genomic enrichment of human exonic sequences through short-fragment libraries |
| Organism |
Homo sapiens |
| Experiment type |
Genome binding/occupancy profiling by high throughput sequencing
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| Summary |
Microarray-based enrichment of selected genomic loci is a powerful method for genome complexity reduction. Since the vast majority of exons in vertebrate genomes are smaller than 150 nt, we have explored the use of short fragment libraries (85-110bp) to achieve higher enrichment specificity by reducing carryover and adverse effects of flanking intronic sequences. These short fragment libraries were enriched for 1.69 Mb of exonic sequences, using custom 244K microarrays, and sequenced using AB/SOLiD. High enrichment specificity (60 – 75%) was obtained at 67-213x average coverage, with 77-92% and 90-98% of targeted regions covered with more than 25% and 10% of the average coverage, respectively. As a more appropriate measure of the evenness of coverage, which is relatively independent of sequencing depth, we introduce the evenness of coverage parameter E. E values up to 75% were achieved. To verify the accuracy of SNP/mutation detection we evaluated 384 known non-reference SNPs in the targeted regions. At ~ 200x average sequence coverage, we were able to survey 96.4% of 1.69 Mb of genomic sequence with only 4.2% false negative calls while 3.6% of targeted regions were marked as unsurveyed. A total of 1197 new variants were detected. Verification revealed only 8 false positive calls, resulting in an overall false positive rate of less than 1 per ~200,000 bp (0.0005%, equivalent to an overall phred score of 55).
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| Overall design |
4 samples + capture design file
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| Contributor(s) |
Mokry M |
| Citation(s) |
20164091 |
| Submission date |
Oct 13, 2009 |
| Last update date |
May 15, 2019 |
| Contact name |
Michal Mokry |
| E-mail(s) |
m.mokry@umcutrecht.nl
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| Organization name |
Wilhelmina Children's Hospital, University Medical Center Utrecht
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| Street address |
Lundlaan 6
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| City |
Utrecht |
| ZIP/Postal code |
3584 EA |
| Country |
Netherlands |
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| Platforms (2) |
| GPL9138 |
AB SOLiD System 2.0 (Homo sapiens) |
| GPL9442 |
AB SOLiD System 3.0 (Homo sapiens) |
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| Samples (4)
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| Relations |
| SRA |
SRP001835 |
| BioProject |
PRJNA121347 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE18542_RAW.tar |
207.6 Mb |
(http)(custom) |
TAR (of BED, TXT) |
| GSE18542_design.bed.gz |
952.7 Kb |
(ftp)(http) |
BED |
SRA Run Selector |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
| Raw data are available in SRA |
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