Premium
Denaturing gradient gel electrophoresis and direct sequencing of PCR amplified genomic DNA: a rapid and reliable diagnostic approach to beta thalassaemia
Author(s) -
Losekoot M.,
Fodde R.,
Harteveld C. L.,
Heeren H. van,
Giordano P. C.,
Bernini L. F.
Publication year - 1990
Publication title -
british journal of haematology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.907
H-Index - 186
eISSN - 1365-2141
pISSN - 0007-1048
DOI - 10.1111/j.1365-2141.1990.tb07883.x
Subject(s) - temperature gradient gel electrophoresis , microbiology and biotechnology , oligonucleotide , polymerase chain reaction , genomic dna , biology , genetics , gene , dna , gel electrophoresis , dna sequencing , 16s ribosomal rna
S ummary The analysis of polymerase chain reaction (PCR)‐amplified β‐globin DNA with allele‐specific oligonucleotide (ASO) probes reveals a very heterogeneous spectrum of β‐thalassaemia in the Netherlands. However about 20% of the β‐thalassaemia mutations cannot be identified with this approach. The combination of specific amplification of certain regions of the β‐globin gene with denaturing gradient gel electrophoresis (DGGE) allowed us to rapidly localize several of these mutations to specific regions of the gene. which were again amplified and directly sequenced. We believe that the combination of DGGE and the direct sequence determination of PCR amplified genomic DNA represents a valid alternative to the‘ASO probes’approach, especially in countries where a very heterogeneous spectrum of β‐thalassaemia mutations occurs.