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Accurate determination of the number of CAG repeats in the Huntington disease gene using a sequence-specific internal DNA standard.
Norge.
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1999 (English)In: Clinical Genetics, ISSN 0009-9163, E-ISSN 1399-0004, Vol. 55, no 3, p. 198-202Article in journal (Refereed) Published
Abstract [en]

We have developed a sequence-specific internal DNA size standard for the accurate determination of the number of CAG repeats in the Huntington disease (HD) gene by cloning key fragments (between 15 and 64 CAG repeats) of the HD gene. These fragments, pooled to produce a sequence-specific DNA ladder, enabled us to observe the true number of CAG repeats directly, with no need for calculations. Comparison of the calculated numbers of CAG repeats in the HD gene using this sequence-specific DNA standard with a commercially available standard (GENESCAN-500 TAMRA) showed that the latter underestimated the number of CAG repeats by three when analyzed by capillary electrophoresis on the ABI 310 Genetic Analyzer (POP4 polymer). In contrast, the use of the same standard overestimated the number of CAG repeats by one when the samples were analyzed by denaturing polyacrylamide electrophoresis on ABI 377 DNA Sequencer (6% denaturing polyacrylamide gel). This suggests that our sequence-specific standard provides greater accuracy for the determination of the true number of CAG repeats in the HD gene than commercially available standards. The sequence-specific standard can be radioactively labeled and successfully replace conventional DNA size standards when analyzing polymerase chain reaction (PCR)-amplified HD alleles by denaturing polyacrylamide electrophoresis.

Place, publisher, year, edition, pages
1999. Vol. 55, no 3, p. 198-202
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Nursing
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URN: urn:nbn:se:esh:diva-7489PubMedID: 10334474OAI: oai:DiVA.org:esh-7489DiVA, id: diva2:1317221
Available from: 2019-05-22 Created: 2019-05-22 Last updated: 2019-05-22Bibliographically approved

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