Multiplex Dideoxynucleotide Sequencing Using Polyadenylated Sequencing Primers Multiplex Dideoxynucleotide Sequencing Using Polyadenylated Sequencing Primers
2013
Abstract
The new approach is used for identification of C282Y (C), H63D (H), and S65C (S) mutations in the HFE gene. In a group of 40 samples we amplified specific parts of HFE exon 2 and exon 4 by multiplex PCR.
Purified amplicons were subsequently used in a multiplex cycle sequencing reaction containing a hyperadenylated reverse primer for exon 2. Adenylation prolonged the migration time of the exon 2 amplicons in the electrophoretic capillary.
This enabled us to obtain separated sequencing data for both the studied amplicons. We found six subjects with the risk of hereditary hemochromatosis (15 %).