Selection of Suitable Reference Genes for Analysis of Salivary Transcriptome in Non-Syndromic Autistic Male Children
Childhood autism is a severe form of complex genetically heterogeneous and behaviorally defined set of neurodevelopmental diseases, collectively termed as autism spectrum disorders (ASD). Reverse transcriptase quantitative real-time PCR (RT-qPCR) is a highly sensitive technique for transcriptome ana...
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doaj-a0ae2c621b6f4352b9900b3a998c5db92020-11-24T21:47:47ZengMDPI AGInternational Journal of Molecular Sciences1422-00672016-10-011710171110.3390/ijms17101711ijms17101711Selection of Suitable Reference Genes for Analysis of Salivary Transcriptome in Non-Syndromic Autistic Male ChildrenYasin Panahi0Fahimeh Salasar Moghaddam1Zahra Ghasemi2Mandana Hadi Jafari3Reza Shervin Badv4Mohamad Reza Eskandari5Mehrdad Pedram6Department of Genetics and Molecular Medicine, School of Medicine, Zanjan University of Medical Sciences (ZUMS), Zanjan 45139-56111, IranDepartment of Medical Biotechnology, School of Medicine, Zanjan University of Medical Sciences (ZUMS), Zanjan 45139-56111, IranDepartment of Medical Biotechnology, School of Medicine, Zanjan University of Medical Sciences (ZUMS), Zanjan 45139-56111, IranDepartment of Genetics and Molecular Medicine, School of Medicine, Zanjan University of Medical Sciences (ZUMS), Zanjan 45139-56111, IranDepartment of Pediatric Neurology, School of Medicine, Tehran University of Medical Sciences (TUMS), Tehran 14176-13151, IranMetrowest CNS Research Center, Natick, MA 01760, USADepartment of Genetics and Molecular Medicine, School of Medicine, Zanjan University of Medical Sciences (ZUMS), Zanjan 45139-56111, IranChildhood autism is a severe form of complex genetically heterogeneous and behaviorally defined set of neurodevelopmental diseases, collectively termed as autism spectrum disorders (ASD). Reverse transcriptase quantitative real-time PCR (RT-qPCR) is a highly sensitive technique for transcriptome analysis, and it has been frequently used in ASD gene expression studies. However, normalization to stably expressed reference gene(s) is necessary to validate any alteration reported at the mRNA level for target genes. The main goal of the present study was to find the most stable reference genes in the salivary transcriptome for RT-qPCR analysis in non-syndromic male childhood autism. Saliva samples were obtained from nine drug naïve non-syndromic male children with autism and also sex-, age-, and location-matched healthy controls using the RNA-stabilizer kit from DNA Genotek. A systematic two-phased measurement of whole saliva mRNA levels for eight common housekeeping genes (HKGs) was carried out by RT-qPCR, and the stability of expression for each candidate gene was analyzed using two specialized algorithms, geNorm and NormFinder, in parallel. Our analysis shows that while the frequently used HKG ACTB is not a suitable reference gene, the combination of GAPDH and YWHAZ could be recommended for normalization of RT-qPCR analysis of salivary transcriptome in non-syndromic autistic male children.http://www.mdpi.com/1422-0067/17/10/1711childhood autismnon-syndromictranscriptomesalivareverse transcriptase quantitative real-time PCR (RT-qPCR)housekeeping genes (HKGs)reference genestability of expressiongeNormNormFinder |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Yasin Panahi Fahimeh Salasar Moghaddam Zahra Ghasemi Mandana Hadi Jafari Reza Shervin Badv Mohamad Reza Eskandari Mehrdad Pedram |
spellingShingle |
Yasin Panahi Fahimeh Salasar Moghaddam Zahra Ghasemi Mandana Hadi Jafari Reza Shervin Badv Mohamad Reza Eskandari Mehrdad Pedram Selection of Suitable Reference Genes for Analysis of Salivary Transcriptome in Non-Syndromic Autistic Male Children International Journal of Molecular Sciences childhood autism non-syndromic transcriptome saliva reverse transcriptase quantitative real-time PCR (RT-qPCR) housekeeping genes (HKGs) reference gene stability of expression geNorm NormFinder |
author_facet |
Yasin Panahi Fahimeh Salasar Moghaddam Zahra Ghasemi Mandana Hadi Jafari Reza Shervin Badv Mohamad Reza Eskandari Mehrdad Pedram |
author_sort |
Yasin Panahi |
title |
Selection of Suitable Reference Genes for Analysis of Salivary Transcriptome in Non-Syndromic Autistic Male Children |
title_short |
Selection of Suitable Reference Genes for Analysis of Salivary Transcriptome in Non-Syndromic Autistic Male Children |
title_full |
Selection of Suitable Reference Genes for Analysis of Salivary Transcriptome in Non-Syndromic Autistic Male Children |
title_fullStr |
Selection of Suitable Reference Genes for Analysis of Salivary Transcriptome in Non-Syndromic Autistic Male Children |
title_full_unstemmed |
Selection of Suitable Reference Genes for Analysis of Salivary Transcriptome in Non-Syndromic Autistic Male Children |
title_sort |
selection of suitable reference genes for analysis of salivary transcriptome in non-syndromic autistic male children |
publisher |
MDPI AG |
series |
International Journal of Molecular Sciences |
issn |
1422-0067 |
publishDate |
2016-10-01 |
description |
Childhood autism is a severe form of complex genetically heterogeneous and behaviorally defined set of neurodevelopmental diseases, collectively termed as autism spectrum disorders (ASD). Reverse transcriptase quantitative real-time PCR (RT-qPCR) is a highly sensitive technique for transcriptome analysis, and it has been frequently used in ASD gene expression studies. However, normalization to stably expressed reference gene(s) is necessary to validate any alteration reported at the mRNA level for target genes. The main goal of the present study was to find the most stable reference genes in the salivary transcriptome for RT-qPCR analysis in non-syndromic male childhood autism. Saliva samples were obtained from nine drug naïve non-syndromic male children with autism and also sex-, age-, and location-matched healthy controls using the RNA-stabilizer kit from DNA Genotek. A systematic two-phased measurement of whole saliva mRNA levels for eight common housekeeping genes (HKGs) was carried out by RT-qPCR, and the stability of expression for each candidate gene was analyzed using two specialized algorithms, geNorm and NormFinder, in parallel. Our analysis shows that while the frequently used HKG ACTB is not a suitable reference gene, the combination of GAPDH and YWHAZ could be recommended for normalization of RT-qPCR analysis of salivary transcriptome in non-syndromic autistic male children. |
topic |
childhood autism non-syndromic transcriptome saliva reverse transcriptase quantitative real-time PCR (RT-qPCR) housekeeping genes (HKGs) reference gene stability of expression geNorm NormFinder |
url |
http://www.mdpi.com/1422-0067/17/10/1711 |
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