Analytical validation of the tag-it high-throughput microsphere-based universal array genotyping platform: application to the multiplex detection of a panel of thrombophilia-associated single-nucleotide polymorphisms.
BACKGROUND
We have developed a novel, microsphere-based universal array platform referred to as the Tag-It platform. This platform is suitable for high-throughput clinical genotyping applications and was used for multiplex analysis of a panel of thrombophilia-associated single-nucleotide polymorphisms (SNPs).


METHODS
Genomic DNA from 132 patients was amplified by multiplex PCR using 6 primer sets, followed by multiplex allele-specific primer extension using 12 universally tagged genotyping primers. The products were then sorted on the Tag-It array and detected by use of the Luminex xMAP system. Genotypes were also determined by sequencing.


RESULTS
Empirical validation of the universal array showed that the highest nonspecific signal was 3.7% of the specific signal. Patient genotypes showed 100% concordance with direct DNA sequencing data for 736 SNP determinations.


CONCLUSIONS
The Tag-It microsphere-based universal array platform is a highly accurate, multiplexed, high-throughput SNP-detection platform.
Bortolin
Susan
S
Tm Bioscience Corporation, Toronto, Ontario, Canada. susanb@tmbioscience.com
Black
Margot
M
Modi
Hemanshu
H
Boszko
Ihor
I
Kobler
Daniel
D
Fieldhouse
Dan
D
Lopes
Eve
E
Lacroix
Jean-Michel
JM
Grimwood
Rebecca
R
Wells
Philip
P
Janeczko
Richard
R
Zastawny
Roman
R
D005838:Genotype; D006801:Humans; D008863:Microspheres; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods; D020641:Polymorphism, Single Nucleotide; D019851:Thrombophilia / Q000235:genetics*