A Novel Electrochemical Biosensor Based on a Double-Signal Technique for d(CAG)_<i>n</i> Trinucleotide Repeats

Electrochemical sensors now play an important role in analysis and detection of nucleic acids. In this work, we present a novel double-signal technique for electrochemically measuring the sequence and length of the d­(CAG)_<i>n</i> repeat. The double-signal technique used an electrochemical molecular beacon (a hairpin DNA labeled with ferrocene), which was directly modified on the surface of a gold electrode, while a reporter probe (a DNA sequence labeled with horseradish peroxidase) was hybridized to the target DNA. First a simple single-signal sensor was characterized in which d­(CAG)_<i>n</i> repeats were detected using a short reporter DNA strand labeled with horseradish peroxidase. To obtain a reliable signal that was dependent on repeat number, a double-signal biosensor was created in which the single strand capture DNA in single-signal sensor was replaced by an electrochemical molecular beacon labeled with ferrocene. When the hairpin DNA hybridized to the target–reporter DNA complex, it opened, resulting in a decreased ferrocene current. Both electrochemical biosensors exhibited high selectivity and sensitivity with low detection limits of 0.21 and 0.15 pM, respectively, for the detection of d­(CAG)_<i>n</i> repeats. The double-signal sensor was more accurate for the determination of repeat length, which was measured from the ratio of signals for HRP and ferrocene (H/F). A linear relationship was found between H/F and the number of repeats (<i>n</i>), H/F = 0.1398<i>n</i> + 9.89788, with a correlation coefficient of 0.974. Only 10 nM of target DNA was required for measurements based on the value of H/F in the double-signal technique. These results indicated that this new double-signal electrochemical sensor provided a reliable method for the analysis of CAG trinucleotide repeats

Table1_A case of malonyl coenzyme A decarboxylase deficiency with novel mutations and literature review.docx

IntroductionMalonyl coenzyme A decarboxylase deficiency is caused by an abnormality in the MLYCD gene. The clinical manifestations of the disease involve multisystem and multiorgan.MethodsWe collected and analyzed a patient's clinical characteristics, genetic chain of evidence and RNA-seq. We use the search term “Malonyl-CoA Decarboxylase Deficiency” on Pubmed to collect cases reported.ResultsWe report a 3-year-old girl who is presented with developmental retardation, myocardial damage and elevated C3DC. High-throughput sequencing identified heterozygous mutation (c.798G>A, p.Q266?) in the patient inherited from her father. The other heterozygous mutation (c.641+5G>C) was found in the patient inherited from her mother. RNA-seq showed that there were 254 differential genes in this child, among which 153 genes were up-regulated and 101 genes were down-regulated. Exon jumping events occurred in exons encoding PRMT2 on the positive chain of chromosome 21, which led to abnormal splicing of PRMT2. (PDiscussionIt is the first report about the locus, adding a new item to the MLYCD mutation library. Developmental retardation and cardiomyopathy are the most common clinical manifestations, with commonly elevated malonate and malonyl carnitine levels in children.</p

Table_2_Sensory organization of balance control in children with vestibular migraine and recurrent vertigo of childhood.DOCX

BackgroundMigraine plays an important role in some subgroups of children with recurrent vertigo. Moreover, the migraine component varies from definite to possibly absent as defined in this spectrum of three disorders—vestibular migraine of childhood (VMC), probable VMC (pVMC), and recurrent vertigo of childhood (RVC). However, studies on the sensory organization of balance control in these three disorders are rare.ObjectiveTo explore the balance control of children with RVC, VMC, and pVMC, when the three sensory systems are challenged.MethodA retrospective analysis was performed on 125 children with VMC (18 female and 15 male; aged 11.64 ± 2.74), pVMC (10 female and eight male; aged 11.78 ± 2.51), and RVC (32 female and 42 male; aged 11.10 ± 2.60). All children in each subtype were divided into groups of children aged ≤ 12 years old and 13–17 years old. Vestibular examination screening and assessment for postural control using the six conditions of the sensory organization test (SOT) were performed. The three primary outcome measures were: equilibrium score (ES), strategy score (SS), and sensory analysis score of the SOT.ResultsEquilibrium score under six different conditions and composite score increased with age (all P-values 0.05). In the children ≤ 12 year-old group, children with VMC had a significantly higher visual preference score than those with pVMC and RVC (P ConclusionCompared with patients at the age of 13–17 years old and with RVC and pVMC (both ≤ 12 years old), children with VMC had a higher degree of reliance on visual signals to maintain their balance and a poorer central integration of peripheral information before reaching 12 years of age. In addition, vision may predominate by weakening vestibular function based on visuo-vestibular interactions. It must be noted that peripheral vestibular examinations could not distinguish the three disease subtypes.</p

Table_1_Sensory organization of balance control in children with vestibular migraine and recurrent vertigo of childhood.DOCX

BackgroundMigraine plays an important role in some subgroups of children with recurrent vertigo. Moreover, the migraine component varies from definite to possibly absent as defined in this spectrum of three disorders—vestibular migraine of childhood (VMC), probable VMC (pVMC), and recurrent vertigo of childhood (RVC). However, studies on the sensory organization of balance control in these three disorders are rare.ObjectiveTo explore the balance control of children with RVC, VMC, and pVMC, when the three sensory systems are challenged.MethodA retrospective analysis was performed on 125 children with VMC (18 female and 15 male; aged 11.64 ± 2.74), pVMC (10 female and eight male; aged 11.78 ± 2.51), and RVC (32 female and 42 male; aged 11.10 ± 2.60). All children in each subtype were divided into groups of children aged ≤ 12 years old and 13–17 years old. Vestibular examination screening and assessment for postural control using the six conditions of the sensory organization test (SOT) were performed. The three primary outcome measures were: equilibrium score (ES), strategy score (SS), and sensory analysis score of the SOT.ResultsEquilibrium score under six different conditions and composite score increased with age (all P-values 0.05). In the children ≤ 12 year-old group, children with VMC had a significantly higher visual preference score than those with pVMC and RVC (P ConclusionCompared with patients at the age of 13–17 years old and with RVC and pVMC (both ≤ 12 years old), children with VMC had a higher degree of reliance on visual signals to maintain their balance and a poorer central integration of peripheral information before reaching 12 years of age. In addition, vision may predominate by weakening vestibular function based on visuo-vestibular interactions. It must be noted that peripheral vestibular examinations could not distinguish the three disease subtypes.</p

Summary of variants.

<p>Summary of variants.</p

SNVs detected in 46 EVA probands.

<p>SNVs detected in 46 EVA probands.</p

Multiple PCR Target Enrichment and Next-Generation Sequencing.

<p>Multiple PCR Target Enrichment and Next-Generation Sequencing.</p

Details of Phenotype of the 46 patients.

<p>Details of Phenotype of the 46 patients.</p

EVA-associated genes and their PCR target regions.

<p>EVA-associated genes and their PCR target regions.</p

Application of a New Genetic Deafness Microarray for Detecting Mutations in the Deaf in China

<div><p>Objective</p><p>The aim of this study was to evaluate the GoldenGate microarray as a diagnostic tool and to elucidate the contribution of the genes on this array to the development of both nonsyndromic and syndromic sensorineural hearing loss in China.</p><p>Methods</p><p>We developed a microarray to detect 240 mutations underlying syndromic and nonsyndromic sensorineural hearing loss. The microarray was then used for analysis of 382 patients with nonsyndromic sensorineural hearing loss (including 15 patients with enlarged vestibular aqueduct syndrome), 21 patients with Waardenburg syndrome, and 60 unrelated controls. Subsequently, we analyzed the sensitivity, specificity, and reproducibility of this new approach after Sanger sequencing-based verification, and also determined the contribution of the genes on this array to the development of distinct hearing disorders.</p><p>Results</p><p>The sensitivity and specificity of the microarray chip were 98.73% and 98.34%, respectively. Genetic defects were identified in 61.26% of the patients with nonsyndromic sensorineural hearing loss, and 9 causative genes were identified. The molecular etiology was confirmed in 19.05% and 46.67% of the patients with Waardenburg syndrome and enlarged vestibular aqueduct syndrome, respectively.</p><p>Conclusion</p><p>Our new mutation-based microarray comprises an accurate and comprehensive genetic tool for the detection of sensorineural hearing loss. This microarray-based detection method could serve as a first-pass screening (before next-generation-sequencing screening) for deafness-causing mutations in China.</p></div