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Infinium Assay for Large-scale SNP Genotyping Applications
Published on: November 19, 2013
Detection of simple mutations and polymorphisms in large genomic regions
E V Sokurenko1, V Tchesnokova, A T Yeung
1Department of Microbiology, University of Washington, Box 357242, Seattle, WA 98195, USA. evs@u.washington.edu
Nucleic Acids Research
|November 20, 2001
Summary
A new technology detects simple nucleotide polymorphisms in total genomic DNA without PCR amplification. This method uses size-fractionation and endonuclease CEL I to identify mutations in bacterial genomes, revealing high stability.
Area of Science:
- Molecular Biology
- Genomics
- Microbiology
Background:
- Detecting simple nucleotide polymorphisms (SNPs) is crucial for understanding genetic variation and disease.
- Current methods often require Polymerase Chain Reaction (PCR) amplification, which can introduce biases.
- A need exists for direct detection methods applicable to large genomic regions.
Purpose of the Study:
- To develop and validate a novel technology for direct detection of SNPs in total genomic DNA.
- To assess the capability of this technology in identifying mutations within bacterial genomes.
- To compare genomic DNA from bacterial clones with divergent phenotypes.
Main Methods:
- Developed a method utilizing preparative size-fractionation of restriction-digested genomic DNA.
- Exploited the mismatch-specific endonuclease CEL I for highly sensitive and specific cleavage of heteroduplex DNA.
- Applied Southern blot hybridization with cosmid DNA libraries and genomic fractions as probes.
Main Results:
- Successfully detected various simple mutations directly in genomic DNA of Pseudomonas aeruginosa, Escherichia coli, and Salmonella isolates.
- Identified a GA insertion in the quorum-sensing regulator gene rhlR in P. aeruginosa.
- Discovered a novel intragenomic polymorphism in rrn operons, indicating high bacterial genome stability.
Conclusions:
- The developed technology enables direct detection of SNPs in hundreds of kilobases of genomic DNA without PCR.
- This method is effective for mutation scanning in bacterial isolates and comparative genomics.
- Bacterial genomes exhibit remarkable stability under natural, non-mutator conditions.
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Sequencing of the human genome has opened up several best-kept secrets of the genome. Scientists have identified thousands of genome variations that exist within a population. These variations can be a single nucleotide or a larger chromosomal variation.
Copy number variations or CNVs are the structural variations that cover more than 1kb of DNA sequence. The single nucleotide polymorphism (SNP), on the other hand, is a single nucleotide change or a point mutation that is found in more than 1%...
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