Detection of the DMD gene deletion using DNA extracted from dried blood spots
Date Issued
2015
Date
2015
Author(s)
Chen, Wei-Chih
Abstract
Background: Duchene muscular dystrophy (DMD) or Becker muscular dystrophy (BMD) are caused by the DMD gene mutation, which is one of the X-linked myopathy that affects mostly males. The prevalence is one in 3600-6000 in male newborns. About 60~70% of these patients can find mutation in the DMD gene, that located at Xp21.2. According to the advance in pharmaceutical development, the progress of patients can be slow down. Examination of the creatine kinase (CK) is what we use in the newborn screening test nowadays; the sensitivity needed to be improved. The average diagnosis ages of DMD is 5 years old. To improve the prognosis, it can achieve by early detection of patients via newborn screening. Purpose: We hope to establish the method to detect DMD gene deletion or duplication by DNA extracted from the dried blood spot (DBS) using multiple ligation-dependent probe amplification (MLPA) as a second tier for those with high CK values. Method: We compared the MLPA result using DNA extracted from traditional method (whole blood) with DNA extracted from DBS using different methods (direct dissolve, filtration, precipitation, DNA kit column, tissue extraction kit). Then, we chose one method from this experiment to establish the standard values. Finally, we tested the clinical samples using traditional methods with our method to check the accuracy of our method. Results: We found that the best way to handle for MLPA was to dissolve DNA in H2O directly. After testing eight clinical samples, we identified four with DMD deletion, one with DMD duplication, and three with normal DMD gene dosage. The results were comparable to traditional results. The accuracy is 100%. Conclusion: We established the MLPA method to detect DMD gene deletion or duplication using DNA from DBS. This method is simple and fast. In the future, it is suitable be the second tier test for high CK in newborn screen.
Subjects
Duchene muscular dystrophy (DMD)
dystrophin
dried blood spot (DBS)
multiple ligation-dependent probe amplification (MLPA)
Type
thesis
File(s)![Thumbnail Image]()
Loading...
Name
ntu-104-P02448006-1.pdf
Size
23.32 KB
Format
Adobe PDF
Checksum
(MD5):8595480d7df62749827f11f98c86c468