repeat units present in a eukaryotic genome would be quite unreasonable. Here we provide an alternative approach for the characterization of a set of internal transcribed spacer sequences found within every rDNA repeat unit by implementing direct sequencing methodology. The prominent allelic variants and their relative amounts characterizing an individual can be described by a single sequencing electropherogram of the mixed amplicon containing the variants present within the genome. We propose a method for rational analysis of heterogeneity of multicopy genes by compiling a profile based on quantification of different sequence variants of the internal transcribed spacers of the freshwater sponge Ephydatia fluviatilis as an example. In addition to using conventional substitution analysis, we have developed a mathematical method, the
to eliminate any spurious nonspecific ampli- cation (higher than six orders of magnitude), fication product. However, the high risk of and the significantly higher reliability of the cross contamination is the main drawback, results compared with conventional PCR. and great care must be taken when perform- There are two different strategies for the ing such PCRs. Several GMO methods based detection of the new amplicon being gener- Table 29.1. GMO specific PCR methods for GMO analysis. cPCR: conventional PCR; RTi-PCR: real- time PCR; dc-PCR: double competitive PCR; qc-PCR: quantitative competitive PCR. Modified from Hernandez et al. (2005) GMO Target sequence Technique Reference Maize event 176 cryIA(b), bla, bar, P-35S/bar, cPCR Ehlers et al., 1997 ivr1
annaabi.ee 
