Abstract
In cystic fibrosis (CF), the most common mutation, delta F508 (a three-base-pair deletion) accounts for ca. 70% of mutations in the worldwide population. The majority of other mutations (more than 350 reported so far to the Genetic Analysis Consortium) have been detected in single cases, thus rendering quite cumbersome a molecular diagnostic approach for the identification of CF chromosomes. As an alternative, linkage analysis based on intragenic polymorphism can be useful for prenatal diagnosis and CF-carrier detection, provided that the heterozygosity of the allelic forms is very high. For this purpose, DNA microsatellites, consisting of two to epta nucleotide repeat clusters, displaying a high degree of polymorphism, are being increasingly used as markers in linkage studies. Two main allelic forms, one hexameric (111 bp) and one heptameric (115 bp), of a tetranucleotide (GATT) repeat polymorphism, at the junction of intron IVS6a and exon 6b, have been amplified by PCR technology. These two alleles can be separated in a 10-20%T polyacrylamide gradient gel and detected by ethidium bromide staining. As an alternate procedure, these two fragments are efficiently separated by capillary zone electrophoresis in a viscous solution of 6%T linear polyacrylamide and detected by their intrinsic absorbance at 254 nm.
| Original language | English |
|---|---|
| Pages (from-to) | 640-643 |
| Number of pages | 4 |
| Journal | Electrophoresis |
| Volume | 15 |
| Issue number | 5 |
| Publication status | Published - 1994 |
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ASJC Scopus subject areas
- Clinical Biochemistry
Cite this
Capillary zone electrophoresis of polymerase chain reaction-amplified DNA fragments in polymer networks : The case of GATT microsatellites in cystic fibrosis. / Gelfi, C.; Orsi, A.; Righetti, P. G.; Brancolini, V.; Cremonesi, L.; Ferrari, M.
In: Electrophoresis, Vol. 15, No. 5, 1994, p. 640-643.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Capillary zone electrophoresis of polymerase chain reaction-amplified DNA fragments in polymer networks
T2 - The case of GATT microsatellites in cystic fibrosis
AU - Gelfi, C.
AU - Orsi, A.
AU - Righetti, P. G.
AU - Brancolini, V.
AU - Cremonesi, L.
AU - Ferrari, M.
PY - 1994
Y1 - 1994
N2 - In cystic fibrosis (CF), the most common mutation, delta F508 (a three-base-pair deletion) accounts for ca. 70% of mutations in the worldwide population. The majority of other mutations (more than 350 reported so far to the Genetic Analysis Consortium) have been detected in single cases, thus rendering quite cumbersome a molecular diagnostic approach for the identification of CF chromosomes. As an alternative, linkage analysis based on intragenic polymorphism can be useful for prenatal diagnosis and CF-carrier detection, provided that the heterozygosity of the allelic forms is very high. For this purpose, DNA microsatellites, consisting of two to epta nucleotide repeat clusters, displaying a high degree of polymorphism, are being increasingly used as markers in linkage studies. Two main allelic forms, one hexameric (111 bp) and one heptameric (115 bp), of a tetranucleotide (GATT) repeat polymorphism, at the junction of intron IVS6a and exon 6b, have been amplified by PCR technology. These two alleles can be separated in a 10-20%T polyacrylamide gradient gel and detected by ethidium bromide staining. As an alternate procedure, these two fragments are efficiently separated by capillary zone electrophoresis in a viscous solution of 6%T linear polyacrylamide and detected by their intrinsic absorbance at 254 nm.
AB - In cystic fibrosis (CF), the most common mutation, delta F508 (a three-base-pair deletion) accounts for ca. 70% of mutations in the worldwide population. The majority of other mutations (more than 350 reported so far to the Genetic Analysis Consortium) have been detected in single cases, thus rendering quite cumbersome a molecular diagnostic approach for the identification of CF chromosomes. As an alternative, linkage analysis based on intragenic polymorphism can be useful for prenatal diagnosis and CF-carrier detection, provided that the heterozygosity of the allelic forms is very high. For this purpose, DNA microsatellites, consisting of two to epta nucleotide repeat clusters, displaying a high degree of polymorphism, are being increasingly used as markers in linkage studies. Two main allelic forms, one hexameric (111 bp) and one heptameric (115 bp), of a tetranucleotide (GATT) repeat polymorphism, at the junction of intron IVS6a and exon 6b, have been amplified by PCR technology. These two alleles can be separated in a 10-20%T polyacrylamide gradient gel and detected by ethidium bromide staining. As an alternate procedure, these two fragments are efficiently separated by capillary zone electrophoresis in a viscous solution of 6%T linear polyacrylamide and detected by their intrinsic absorbance at 254 nm.
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M3 - Article
C2 - 7925242
AN - SCOPUS:0028221422
VL - 15
SP - 640
EP - 643
JO - Electrophoresis
JF - Electrophoresis
SN - 0173-0835
IS - 5
ER -