Science Supplement: Using RNA sequencing to reveal hidden mutations that cause genetic diseases
April 24, 2017 Source: Bio Valley
Window._bd_share_config={ "common":{ "bdSnsKey":{ },"bdText":"","bdMini":"2","bdMiniList":false,"bdPic":"","bdStyle":" 0","bdSize":"16"},"share":{ }};with(document)0[(getElementsByTagName('head')[0]||body).appendChild(createElement('script')) .src='http://bdimg.share.baidu.com/static/api/js/share.js?v=89860593.js?cdnversion='+~(-new Date()/36e5)];In a new study, RNA sequencing (RNA-seq) of affected tissues can be used to identify mutations that may cause Mendelian genetic disease, according to researchers from research institutions such as the Broad Institute in the United States. . Under the leadership of Dr. Daniel MacArthur, a researcher at the Broad Institute, a team of researchers performed RNA sequencing of skeletal muscle biopsies from patients with rare undiagnosed genetic diseases, revealing that new DNA sequencing is not so easy to identify. Pathogenic mutations, such as pathogenic splice site variants. This study is by far the largest transcriptome sequencing in a group of patients with undiagnosed disease in order to identify previously unknown genetic disease-associated mutations. The results of the study were published in the April 19, 2017 issue of Science Translational Medicine, entitled "Improving genetic diagnosis in Mendelian disease with transcriptome sequencing".
Tuuli Lappalainen, a researcher at the New York Genomics Center in the United States (not involved in the study) said, “This is a very good study that is excellent in confirming that RNA sequencing can be used to discover and describe significant human disease mutations.â€
Jeffrey Barrett, a researcher at the Sanger Institute at the University of Cambridge's Welkom Foundation (not participating in the study), told Scientists that "there has been tremendous progress in how we can use DNA sequencing to diagnose rare genetic diseases. This method still does not identify significant mutations in approximately 60% of cases. This new paper is a good demonstration of the progress made in using RNA sequencing to find these mutations."
To validate this transcriptome sequencing approach, the researchers first performed RNA sequencing of skeletal muscle samples from 13 previously diagnosed patients with skeletal muscle genetic disease. They compared these RNA sequences obtained with those from the RNA sequencing data set of 180 healthy people participating in the Genotype-Tissue Expression Consortium (GTEx). They then performed the same analysis of skeletal muscle samples from 50 patients with undiagnosed skeletal muscle genetic disease.
MacArthur told Scientists that "the benefit of using RNA sequencing to diagnose genetic diseases is that we can now directly study the effects of DNA changes on gene expression and RNA splicing."
These researchers were able to identify pathogenic gene mutations among 17 patients with an overall reduction rate of 35%. The identified mutations include new splice site variants located deep in the non-coding region of the gene and disrupted splice sites that are not located near the exon. For example, in four patients, they were able to identify pathogenic mutations that were not previously detected in the dystrophin gene in patients with Duchenne muscular dystrophy. This analysis revealed a previously unknown severe collagen VI-associated myopathy intron mutation in four patients who were negative for standard DNA sequencing of the collagen VI gene. They then identified this mutation in another 27 patients who were part of a larger population of collagen VI-associated myopathy patients.
According to MacArthur, RNA sequencing adds a way to identify pathogenic mutations, increasing the rate of diagnosis to more than half. He noted that the pathogenic mutations identified in the current study affect genes that are known to cause disease when a mutation occurs. These researchers found more pathogenic genetic mutations associated with the disease.
The key to this transcriptome technology is the use of tissues that express this recognized genetic mutation. Barrett notes, "To prove that this is a good diagnostic method for genetic diseases, these researchers use the muscle tissue they know to be available for this genetic mutation to conduct research."
However, when it is not possible to obtain diseased tissue (such as tissues affected by brain diseases), MacArthur and colleagues are now exploring another way to obtain a patient's skin or blood samples and convert them into induced pluripotent stem cells (iPSC). These iPSCs are then able to differentiate into neurons or other cell types of interest.
Barrett said that for approximately 50% to 60% of genetically ill patients with unknown causes, it may be beneficial to accurately find disease-associated mutations in non-coding regions of the human genome. He said, "When we study the genetic properties of many different human diseases, there are also diseases that are not classical Mendelian genetic diseases (caused by a single mutation) but involve multiple genes." (BioValley Bioon.com )
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