David Tabb
Faculty Member
Last active: 6/26/2014

MS for identification of single nucleotide polymorphisms and MS/MS for discrimination of isomeric PCR products.

Krahmer MT, Walters JJ, Fox KF, Fox A, Creek KE, Pirisi L, Wunschel DS, Smith RD, Tabb DL, Yates JR
Anal Chem. 2000 72 (17): 4033-40

PMID: 10994962 · DOI:10.1021/ac000142b

ESI (electrospray ionization) MS and tandem mass spectrometry (MS/MS) were used for the analysis of single nucleotide polymorphisms (SNPs) and more complex genetic variations. Double-stranded (ds) PCR products were studied. PCR products of the proline [5'-x(G17)-x(C38)x-3'] and arginine variants [(5'-x(Gl7)-x(G38)x-3'] of the p53 gene are distinguished by an SNP (cytosine or guanine) and were discriminated using both quadrupole and quadrupole ion trap MS analysis. A 69 bp arginine mutant PCR product [5'-x(C17)-x(G38)x-3'] with a negating switch has the same mass as the proline variant but was readily distinguishable on ion trap MS/MS analysis; fragments containing the mutation site, but not the polymorphism, were identified. The 69 bp PCR products were restriction-enzyme-digested, to create 43 bp fragments. ESI quadrupole ion trap MS/MS analysis of the 43 bp product-ion spectra readily demonstrated both polymorphism and negating switch sites. MS and MS/MS are powerful and complementary techniques for analysis of DNA. MS can readily distinguish SNPs but MS/MS is required to differentiate isomeric PCR products (same nucleotide composition but different sequence).

MeSH Terms (3)

Mass Spectrometry Polymerase Chain Reaction Polymorphism, Genetic

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