2008 Barcelona - European Society of Human Genetics

Genomics, technology, bioinformatics 0
that ULS-labelling yielded satisfying degrees of labelling (DoL~1 .5%),
while the ones for random priming were ~0 .2% . Reproducibility of the
analysis of these samples on BAC arrays was better as well (R2=0 .96
for ULS vs R2=0 .88 for random priming) .
In addition, aCGH applications involve substantial amounts of genomic
DNA, while the amount of DNA available is often limited, requiring
whole-genome amplification (WGA). Recently, Qiagen’s REPLI-g
has been optimized for degraded samples . Therefore, we compared
BAC aCGH experiments using 100ng of FFPE DNA, labelled using
either direct random priming or a combination of REPLI-g and ULS
or REPLI-g and random priming . Correlation with data obtained using
conventional microgram amounts of input DNA data was best for the
REPLI-g/ULS combination (R2=0 .84 vs 0 .70 for random priming), as
was reproducibility .
In conclusion, ULS allows aCGH analysis of archival DNA, while minimizing
labelling-induced bias .
P08.31
Functional SNP candidates identified by genome wide ChIP
analyses
A. Ameur 1 , A. Rada-Iglesias 2,3 , O. Wallerman 2 , M. Motallebipour 2 , S. Enroth 1 , J.
Komorowski 1,4 , C. Wadelius 2 ;
1 The Linnaeus Centre for Bioinformatics, Uppsala, Sweden, 2 Rudbeck Laboratory,
Uppsala, Sweden, 3 Current address: The Linnaeus Centre for Bioinformatics,
Uppsala, Sweden, 4 Interdisciplinary Centre for Mathematical and Computer
Modelling, Warsaw, Poland.
Disease-associated SNPs identified in large association studies are
often located in non-coding regions and it is possible that some SNPs
affect the interaction between transcription factors and DNA . In HepG2
cells we mapped binding sites for the transcription factors USF1 and
USF2 along the genome using ChIP-chip . Using the novel BCRANK
algorithm, we detected 1754 binding sites for USF1 with the predicted
binding motif . In dbSNP, we found 140 candidate functional SNPs in
these motifs . Eight of the SNPs were heterozygous in at least one of
four human cell samples . In 5 of 6 cases where the SNP was inside the
core predicted USF1 binding motif CACGTGAC, ChIP and sequence
analysis of the enriched DNA showed that USF1 was, with statistical
significance bound to the allele containing the USF1 consensus and
less to the other. We detected significant allelic differences for USF2,
POLR2A and H3K27me3 . As a negative control, we analyzed 2 cases
where the SNP was located just outside USF1 core sequence and saw
no differences for any investigated factor . By high resolution genotyping
in the HepG2 genome we have a dense map of potential functional
SNPs . We have mapped additional factors along the genome using
ChIP-sequencing, identified individual binding motifs containing hundreds
of SNPs. Furthermore, we found significant enrichment of one
allele at heterozygous positions as indication of allele specific DNAprotein
interactions. Thus, individual binding sites can be defined at
base pair resolution and large numbers of candidate functional SNPs
can be predicted in genome wide experiments .
P08.32
TaqMan® Express: A format for easy mRNA quantification using
pre-plated taqman Gene Expression Assays
K. Poulter, F. Chan, J. Rappleye, P. Hegerich, D. N. Keys, K. Y. Lee, G. Marcus,
S. Koepf, C. Chen;
Applied Biosystems, Foster City, CA, United States.
TaqMan® Express Plates contain up to 96 pre-spotted and dried Taq-
Man Gene Expression Assays on an optical 96-well plate . Pre-plated
assays simplify, accelerate and help error-proof mRNA quantification
experiments . This is especially important when there is more than one
operator in a lab, when a study is being done across multiple labs, or
when a study is extended over a period of time . The assays in Taqman®
Express plates are user-selected from a catalogue of >50,000
Taqman assays targeting human, mouse, rat, Rhesus and dog genes .
Here, we show that TaqMan assays which are aliquoted and dried for
storage perform comparably to standard wet assays aliquoted immediately
before use . Dynamic range for dried assays showed >6 logs
of linearity, detection of 2-fold discrimination with 99.7% confidence
was demonstrated, and limit of detection was