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Detection of p53 Gene in Breast Cancer by Denaturing Gradient Gel Electrophoresis and the DCode System

on using a 25 to 65% gradient. Therefore a gradient of 45 to 60% was used for parallel gel analysis.

DNA from 34 cases was found to have a mutation on perpendicular gels and in 33 cases on parallel gels. The sample missed showed a mutation of exon 7 on perpendicular gels but not on parallel gels. A possible explanation for this could be that the parallel gel gradient of 45 to 60% was too wide for that particular sample. A narrow gradient may be required to identify the mutation of exon 7 on parallel gels.

We chose to use parallel gradient method because it provides high throughput analysis. The detection of mutation was unclear in some cases where the rate of mutated allele was low or the genomic DNA was extracted from paraffin embedded materials. In such cases we used SYBR Green I instead of ethidium bromide to stain the gels. This improved sensitivity and enabled us to see the bands clearly.

These experiments indicate that DGGE is a powerful technique to screen mutations. The DCode system is easy-to-use and reliable, two important factors when scanning numerous mutations.


References

1. Foddel, R. and Losekoot, M., Human Mutation, 3, 8394 (1994).

2. Anderson, T. I. and Boreson, A-L., Diag. Mol. Patho., 4, 203211 (1995).


The Polymerase Chain Reaction (PCR) process is covered by patents owned by Hoffmann-LaRoche. Use of the PCR process requires a license.


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