Surface amplification of invasive cleavage products.
A major focus of current efforts in genomics is to elucidate the genetic variations extent within the human population, and to study the effects of these variations upon the human system. The most common type of genetic variations are the single nucleotide polymorphisms (SNPs), which occur every 500-1000 nt in the genome. Large-scale population association studies to study the biological or medical significance of such variations may require the analysis of hundreds of thousands of SNPs on thousands of individuals. We are pursuing development of an approach to large-scale SNP analysis that combines the specificity of invasive cleavage reactions with the parallelism of high density DNA arrays. A surface-immobilized probe oligonucleotide is specifically cleaved in the presence of a complementary target sequence in unamplified human genomic DNA, yielding a 5' phosphate group. High sensitivity detection of this reaction product on the surface is achieved by the use of rolling circle amplification, with an approximate concentration detection limit of 10 fM target DNA. This combination of very specific surface cleavage and highly sensitive surface detection will make possible the rapid and parallel analysis of genetic variations across large populations.
Chen
Yan
Y
Department of Chemistry, University of Wisconsin-Madison, 1101 University Avenue, Madison, Wisconsin 53706-1396, USA.
Shortreed
Michael R
MR
Peelen
Dora
D
Lu
Manchun
M
Smith
Lloyd M
LM
D004247:DNA / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000737:chemistry / Q000378:metabolism; D005453:Fluorescence; D031541:Fluorescence Resonance Energy Transfer; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D020641:Polymorphism, Single Nucleotide
D004247:DNA; D003850:Deoxyribonuclease I