DBS Screening for Glycogen Storage Disease Type 1a: Detection of c.648G>T Mutation in <i>G6PC</i> by Combination of Modified Competitive Oligonucleotide Priming-PCR and Melting Curve Analysis
Glycogen storage disease type Ia (GSDIa) is an autosomal recessive disorder caused by glucose-6-phosphatase (G6PC) deficiency. GSDIa causes not only life-threatening hypoglycemia in infancy, but also hepatocellular adenoma as a long-term complication. Hepatocellular adenoma may undergo malignant tra...
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MDPI AG
2021-11-01
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| Series: | International Journal of Neonatal Screening |
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| Online Access: | https://www.mdpi.com/2409-515X/7/4/79 |
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| author | Emma Tabe Eko Niba Yogik Onky Silvana Wijaya Hiroyuki Awano Naoko Taniguchi Yasuhiro Takeshima Hisahide Nishio Masakazu Shinohara |
| author_facet | Emma Tabe Eko Niba Yogik Onky Silvana Wijaya Hiroyuki Awano Naoko Taniguchi Yasuhiro Takeshima Hisahide Nishio Masakazu Shinohara |
| author_sort | Emma Tabe Eko Niba |
| collection | DOAJ |
| description | Glycogen storage disease type Ia (GSDIa) is an autosomal recessive disorder caused by glucose-6-phosphatase (G6PC) deficiency. GSDIa causes not only life-threatening hypoglycemia in infancy, but also hepatocellular adenoma as a long-term complication. Hepatocellular adenoma may undergo malignant transformation to hepatocellular carcinoma. New treatment approaches are keenly anticipated for the prevention of hepatic tumors. Gene replacement therapy (GRT) is a promising approach, although early treatment in infancy is essential for its safety and efficiency. Thus, GRT requires screening systems for early disease detection. In this study, we developed a screening system for GSDIa using dried blood spots (DBS) on filter paper, which can detect the most common causative mutation in the East-Asian population, c.648G>T in the <i>G6PC</i> gene. Our system consisted of nested PCR analysis with modified competitive oligonucleotide priming (mCOP)-PCR in the second round and melting curve analysis of the amplified products. Here, we tested 54 DBS samples from 50 c.648G (wild type) controls and four c.648T (mutant) patients. This system, using DBS samples, specifically amplified and clearly detected wild-type and mutant alleles from controls and patients, respectively. In conclusion, our system will be applicable to newborn screening for GSDIa in the real world. |
| first_indexed | 2024-03-10T03:53:55Z |
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| language | English |
| last_indexed | 2024-03-10T03:53:55Z |
| publishDate | 2021-11-01 |
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| series | International Journal of Neonatal Screening |
| spelling | doaj.art-aca0a38257e740838b0bffab6192e7072023-11-23T08:50:12ZengMDPI AGInternational Journal of Neonatal Screening2409-515X2021-11-01747910.3390/ijns7040079DBS Screening for Glycogen Storage Disease Type 1a: Detection of c.648G>T Mutation in <i>G6PC</i> by Combination of Modified Competitive Oligonucleotide Priming-PCR and Melting Curve AnalysisEmma Tabe Eko Niba0Yogik Onky Silvana Wijaya1Hiroyuki Awano2Naoko Taniguchi3Yasuhiro Takeshima4Hisahide Nishio5Masakazu Shinohara6Department of Community Medicine and Social Healthcare Science, Kobe University Graduate School of Medicine, 7-5-1 Kusunoki-cho, Kobe 650-0017, JapanDepartment of Community Medicine and Social Healthcare Science, Kobe University Graduate School of Medicine, 7-5-1 Kusunoki-cho, Kobe 650-0017, JapanDepartment of Pediatrics, Kobe University Graduate School of Medicine, 7-5-1 Kusunoki-cho, Kobe 650-0017, JapanDepartment of Pediatrics, Hyogo College of Medicine, 1-1 Mukogawa-cho, Nishinomiya 663-8501, JapanDepartment of Pediatrics, Hyogo College of Medicine, 1-1 Mukogawa-cho, Nishinomiya 663-8501, JapanDepartment of Community Medicine and Social Healthcare Science, Kobe University Graduate School of Medicine, 7-5-1 Kusunoki-cho, Kobe 650-0017, JapanDepartment of Community Medicine and Social Healthcare Science, Kobe University Graduate School of Medicine, 7-5-1 Kusunoki-cho, Kobe 650-0017, JapanGlycogen storage disease type Ia (GSDIa) is an autosomal recessive disorder caused by glucose-6-phosphatase (G6PC) deficiency. GSDIa causes not only life-threatening hypoglycemia in infancy, but also hepatocellular adenoma as a long-term complication. Hepatocellular adenoma may undergo malignant transformation to hepatocellular carcinoma. New treatment approaches are keenly anticipated for the prevention of hepatic tumors. Gene replacement therapy (GRT) is a promising approach, although early treatment in infancy is essential for its safety and efficiency. Thus, GRT requires screening systems for early disease detection. In this study, we developed a screening system for GSDIa using dried blood spots (DBS) on filter paper, which can detect the most common causative mutation in the East-Asian population, c.648G>T in the <i>G6PC</i> gene. Our system consisted of nested PCR analysis with modified competitive oligonucleotide priming (mCOP)-PCR in the second round and melting curve analysis of the amplified products. Here, we tested 54 DBS samples from 50 c.648G (wild type) controls and four c.648T (mutant) patients. This system, using DBS samples, specifically amplified and clearly detected wild-type and mutant alleles from controls and patients, respectively. In conclusion, our system will be applicable to newborn screening for GSDIa in the real world.https://www.mdpi.com/2409-515X/7/4/79glycogen storage disease type 1adried blood spotallele-specific PCRmCOP-PCRmelting curve |
| spellingShingle | Emma Tabe Eko Niba Yogik Onky Silvana Wijaya Hiroyuki Awano Naoko Taniguchi Yasuhiro Takeshima Hisahide Nishio Masakazu Shinohara DBS Screening for Glycogen Storage Disease Type 1a: Detection of c.648G>T Mutation in <i>G6PC</i> by Combination of Modified Competitive Oligonucleotide Priming-PCR and Melting Curve Analysis International Journal of Neonatal Screening glycogen storage disease type 1a dried blood spot allele-specific PCR mCOP-PCR melting curve |
| title | DBS Screening for Glycogen Storage Disease Type 1a: Detection of c.648G>T Mutation in <i>G6PC</i> by Combination of Modified Competitive Oligonucleotide Priming-PCR and Melting Curve Analysis |
| title_full | DBS Screening for Glycogen Storage Disease Type 1a: Detection of c.648G>T Mutation in <i>G6PC</i> by Combination of Modified Competitive Oligonucleotide Priming-PCR and Melting Curve Analysis |
| title_fullStr | DBS Screening for Glycogen Storage Disease Type 1a: Detection of c.648G>T Mutation in <i>G6PC</i> by Combination of Modified Competitive Oligonucleotide Priming-PCR and Melting Curve Analysis |
| title_full_unstemmed | DBS Screening for Glycogen Storage Disease Type 1a: Detection of c.648G>T Mutation in <i>G6PC</i> by Combination of Modified Competitive Oligonucleotide Priming-PCR and Melting Curve Analysis |
| title_short | DBS Screening for Glycogen Storage Disease Type 1a: Detection of c.648G>T Mutation in <i>G6PC</i> by Combination of Modified Competitive Oligonucleotide Priming-PCR and Melting Curve Analysis |
| title_sort | dbs screening for glycogen storage disease type 1a detection of c 648g t mutation in i g6pc i by combination of modified competitive oligonucleotide priming pcr and melting curve analysis |
| topic | glycogen storage disease type 1a dried blood spot allele-specific PCR mCOP-PCR melting curve |
| url | https://www.mdpi.com/2409-515X/7/4/79 |
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