Sensitive Detection of Polyalanine Expansions in PHOX2B by Polymerase Chain Reaction Using Bisulfite-Converted DNA

Horiuchi, Hidekazu; Sasaki, Ayako; Osawa, Motoki; Kijima, Kazuki; Ino, Yukiko; Matoba, Ryoji; Hayasaka, Kiyoshi

doi:10.1016/s1525-1578(10)60598-3

Cited by 18 publications

(12 citation statements)

References 16 publications

(24 reference statements)

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“…Because the melting temperatures of C-rich (and CG-rich) DNA regions are tremendously reduced after the conversion of unmethylated cytosines to uracils with bisulfite deamination, this technique has been successfully applied in tests of expansions of repeats with high GC contents for both methylated and unmethylated repeats (18,19,24,25 ). Thus, this technique also permits testing for repeat-expansion traits such as fragile X syndrome (MIM 309550) (19 ), myoclonus epilepsy, type Unverricht-Lundborg repeat expansion in CTSB [cystatin B (stefin B)] (MIM 254800) (18 ), and testing for congenital central hyperventilation syndrome [PHOX2B (paired-like homeobox 2b) polyalanine expansion; MIM 209880] (25 ).…”

Section: Discussionmentioning

confidence: 99%

PCR-Based Analysis of Differentially Methylated Regions of GNAS Enables Convenient Diagnostic Testing of Pseudohypoparathyroidism Type Ib

et al. 2008

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BACKGROUND: Pseudohypoparathyroidism type Ib (PHPIb) is characterized by parathyroid hormone (PTH) resistance, which can lead to hypocalcemia, hyperphosphatemia, and increased serum PTH. The disorder is caused by mutations in regulatory regions of the GNAS gene (GNAS complex locus) that lead to interferences in the methylation status of alternative GNAS promoters, such as exon A/B, NESP55, and XL␣-s. PHPIb comprises disorders that show distinctive changes in methylation status but share the same clinical phenotype: (a) loss of methylation only at exon A/B of the GNAS gene and involving no other obvious epigenetic abnormalities [e.g., those caused by heterozygous microdeletions in the STX16 (syntaxin 16) region and found in many patients with autosomal dominant (AD) PHPIb]; (b) methylation abnormalities at several differentially methylated regions (DMRs), which are observed in most patients with sporadic PHPIb and some families with AD PHPIb.

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Section: Discussionmentioning

confidence: 99%

PCR-Based Analysis of Differentially Methylated Regions of GNAS Enables Convenient Diagnostic Testing of Pseudohypoparathyroidism Type Ib

et al. 2008

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“…This has represented a considerable problem for diagnostic aims because patients heterozygous for expanded allele appear to be homozygous for the wild-type allele. To overcome this problem, it has recently been reported that amplification of both PHOX2B alleles can occur through the deamination of template DNA, 16 and we used a kit for GC-rich DNA sequences provided by Roche. 13 This sequence was therefore the ideal final challenge to test the combination of the three reagents.…”

Section: Amplification Of Phox2b Exonmentioning

confidence: 99%

Betaine, Dimethyl Sulfoxide, and 7-Deaza-dGTP, a Powerful Mixture for Amplification of GC-Rich DNA Sequences

Musso

Bocciardi

Parodi

et al. 2006

The Journal of Molecular Diagnostics

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Currently, polymerase chain reaction is the most used technique in many laboratories for either diagnostic or molecular biology purposes. Despite the large number of DNA sequences that can be easily analyzed, some GC-rich sequences are refractory to amplification due to the formation of secondary intramolecular structures. To overcome this problem, several molecules have been described to improve polymerization. Here we show that a combination of three additives-betaine, dimethyl sulfoxide, and 7-deaza-dGTP-was essential to achieve amplification of DNA sequences of three disease genes showing a GC content ranging from 67 to 79%.

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“…Mutations or polymorphic changes associated with SIDS have been reported in the following: complements (C4A, C4B) (5,6), IL-10 (7), mitochondrial DNA (MTTL1, MTND1), serotonin transporter (5-HTT) (8,9), a gene associated with sex differentiation (TSPYL) (10), and cardiac ion channels (KCNQ1, KCNH2, SCN5A). We have previously investigated the relationship between SIDS and congenital central hypoventilation syndrome (CCHS); however, we failed to find any mutations other than three single nucleotide polymorphisms (SNPs) of PHOX2B, a gene responsible for CCHS, in 48 Japanese SIDS victims (11,12).…”

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confidence: 99%

Cardiac Ion Channel Gene Mutations in Sudden Infant Death Syndrome

et al. 2008

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Sudden infant death syndrome (SIDS) is multifactorial and may result from the interaction of a number of environmental, genetic, and developmental factors. We studied three major genes causing long QT syndrome in 42 Japanese SIDS victims and found five mutations, KCNQ1-K598R, KCNH2-T895M, SCN5A-F532C, SCN5A-G1084S, and SCN5A-F1705S, in four cases; one case had both KCNH2-T895M and SCN5A-G1084S. All mutations were novel except for SCN5A-F532C, which was previously detected in an arrhythmic patient. Heterologous expression study revealed significant changes in channel properties of KCNH2-T895M, SCN5A-G1084S, and SCN5A-F1705S, but did not in KCNQ1-K598R and SCN5A-F532C. Our data suggests that nearly 10% of SIDS victims in Japan have mutations of the cardiac ion channel genes similar to in other countries.

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Sensitive Detection of Polyalanine Expansions in PHOX2B by Polymerase Chain Reaction Using Bisulfite-Converted DNA

Cited by 18 publications

References 16 publications

PCR-Based Analysis of Differentially Methylated Regions of GNAS Enables Convenient Diagnostic Testing of Pseudohypoparathyroidism Type Ib

PCR-Based Analysis of Differentially Methylated Regions of GNAS Enables Convenient Diagnostic Testing of Pseudohypoparathyroidism Type Ib

Betaine, Dimethyl Sulfoxide, and 7-Deaza-dGTP, a Powerful Mixture for Amplification of GC-Rich DNA Sequences

Cardiac Ion Channel Gene Mutations in Sudden Infant Death Syndrome

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