Bases Added to the Primer

The base or bases added to the primer are identified by comparing tbe mass differences between tbe imextended genotyping primer and tbe corresponding primer extensions with tbe calculated masses of the dideoxynucleotide base residues (Table 1). High mass accuracy is not required, and calibration of tbe mass 

MODIFICATION OF THE SNP TYPING ASSAY TO SUPPORT ALLELE FREQUENCY DETERMINATION 

Although MALDI-TOF is poorly quantitative in the absolute sense, it is good for relative quantitation. That is, when comparing peak areas of MALDI peaks of DNA of about the same size and base composition, the peak areas will accurately reflect the relative proportions of these substances. For example, in the case of a primer extended from a heterozygote target, the two different bases added to the primer will normally produce equal peak areas within a relative factor of +/- 10%. 

Cargill M, Alshuler D, Ireland J, Sklar P, Ardlie K, Patil N, Lane CR, Lim EP, Kalyanaraman N, Nemesh j, Ziaugra L, Friedland L, Rolfe A, Warrington J, Lipshutz R, Daley GQ, Lander ES. Nature Genetics 22 231, 1999 

van Rensburg SJ, Potocnik FCV, deVilliers JNP, Kotze Ml, Taljaard UP. Annals of New York Academy of Science 903 200, 2000 

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MALDI-TOF mass spectrometry is very high in resolution, the available degree of multiplexing is much higher than using fluorescent detection, which is typically limit due to a great deal of spectral overlap between different dyes. Allele determinations by mass spectrometry are nearly artefact-free, because the process measures the intrinsic mass of the bases added to the primer. These mass measurements are not affected by secondary considerations, such as the conformation oftheDNA, which can confound gel-based techniques. 

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