The first time using next generation sequencing in the research of Vietnamese patients with autism spectrum disorders - Identification of two novel missense mutations of RYR3 gene related to transportation of Ca2+ ions

Autism is a disorder of the pervasive developmental disorder (Autism spectrum disorder). Early onset in the first three years of life, affecting the development of children in three main areas such as social interaction, language and behavior. In the United States, the incidence of autism has risen ten-fold since the 1980s, with an estimated one six-eighth of children diagnosed with autism. The number of boys diagnosed with autism was shown to be higher than girls. In Vietnam, increasing number of children with autism has created a great burden and challenge for families and society. Autism has been proven to be a disease caused by both genetic and environmental effects. So far, more than 100 genes related to autism have been identified.

Whole exome sequencing (WES) is an application of next generation sequencing technology used to identify variants across all coding regions of the genome. WES has been increasingly used in the studies of serious diseases, difficult diagnosis, and undetectable causes such as cancer, cardiovascular, neurological diseases etc... Recently, gene sequencing technology has been using worldwide for the diagnosis and treatment of autism.


Recognizing the importance of this technology, Vietnam Academy of Science and Technology has assigned the Institute of Genome Research to implement the project "Whole-exome sequencing of patients with autism in Vietnam ". Receiving the task, the research group headed by Assoc. Prof. Nguyen Huy Hoang has conducted the project for two years (2015-2016) with research contents including screening and selection of patients with autism in Vietnam, whole exome sequencing of the patients with autism by the next generation Illumina gene sequencing machine with a minimum 30X coverage from total DNA, analysis and evaluation of the genetic variances associated with autism.


Until now, the research group has collected seven sets of records and samples of the patients with autism and their family members in Vietnam. Whole-exome sequencing of the seven patients with autism has been done by Illumina next-generation sequencing method. A process of exome sequencing and bioinformatic analysis of the patients with autism has been constructed. A data set of gene mutations and genetic variants in coding regions related to autism in the seven patients has been documented. The data of 285 genetic variants of 101 genes of seven patients related to autism has been collected after comparing with the data of genes sensitive to ASD by Gene Tests and ApolloGen. Sixteen genes sensitive to the ASD relating to ion channel in seven patients have been identified as: CACNA1C, CACNA1D, CACNA1E, CACNA1F, CACNA1G, CACNA1H and CACNA1I. Data of three pathways of the genes (66 genes) related to the characteristic manifestations of the studied patients (repeated habit, flavor recognizability, and the brain development), of which five characteristic genes were shown to share interactions in three pathways as: AFF2, NTRK3, CAMK2B, ATP2B2, and GNAO1.


In particular, two new heterozygous missense mutations, c.332 T>C and c.9142 C>T have been identified by analyzing the genes involved in the abnormalities of internal calcium ion balance in the nervous development in RYR3 gene. The mutation c.332 T>C was found to be inherited from the father whereas the remaining mutation c.9142 C>T from the mother. The mutation c.332 T>C was shown to occur in exon 4, where T was replaced by C at position 332 in the cDNA, resulting in a replacement of the Leu (L) amino acid by Pro (P) at position 111 in RYR3 protein. The mutation c.9142 C>T was detected in exon 65, where C was replaced by T at position 9142, resulting in an alteration of an amino acid at position 3048 where Arg (R) was replaced by Cys (C) in RYR3 protein. These two mutant points were verified by sequencing analysis of the patient’s mother and father on the ABI 3500 genetic analyzer.




Sanger sequencing of exon 4 and exon 65 and the patient family pedigree


In addition to the research outcomes, the project has contributed to the training of a doctor. A part of the results has been reported on a national journal, Journal of Biotechnology and on an international journal, Genes & Genomics (


On February 21, 2017, the project outcomes received the excellent grade from the VAST acceptance committee. This could be considered as the first research in Vietnam in the field of research on the genetics of autism by next generation gene sequencing method. The study of the whole exome of this patient is fully in accordance with the trend of individual genetic research in the world. The result of this project provides an additional knowledge to the view of autism for genetic researchers and doctors, helping to give the best diagnosis and therapy to the patients as well as appropriate genetic counsels to the patients’ families.


Translated by Tuyet Nhung
Link to Vietnamese Version 

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