| Objectives Autism spectrum disorder(ASD)is a common neurodevelopmental disorder characterized by persistent deficits in social communication and social interaction across multiple contexts and restricted,repetitive patterns of behavior,interests,or activities.At present,ASD lacks of early screening method,which affects the early diagnosis,treatment and intervention of disease and the prognosis of children.The epigenetic mechanism can affect the expression of neural development gene network and lead to ASD phenotype.miRNA,one of the most important mechanisms of epigenetic,characterized of extracellular stability,is a biological index for early screening of many diseases.Methods In August 2018,5 ASD children were recruited from Qingdao women’s and children’s Hospital Affiliated to Qingdao University,and 5 normal children matched with the case group in age and gender were selected as controls in the same period.The venous blood of upper arm of all subjects was collected,and the serum was frozen and stored within 4 hours.The total RNA was extracted by Trizol method and the sample quality was evaluated.Send qualified quality control samples to the testing company for subsequent sample processing and high-throughput sequencing.The original sequence obtained from superior sequencing was filtered,the target sequence was classified and annotated,and miRNAs with different expression levels between the two groups were screened.Using the miwalk3.0 website to predict the target genes that may be regulated by differentially expressed mi RNA in multiple databases.The target gene was enriched with go by David website,and then analyzed with KEGG pathway by kobas3.0 website.Results1.The quality of total RNA extracted from 10 samples of the two groups met the requirements of the follow-up experiment,and gene sequencing could be carried out.2.The raw tag obtained by high-throughput sequencing was filtered and the length of the target tag RNA was concentrated in 21-23 nt.3.The target tag was annotated by small RNA classification,and miRNA was the main small RNA in all samples.4.Quantitative analysis of miRNA expression was performed.Compared with the control group,there were 26 miRNAs expressed differently in the case group(P<0.001;FC vaule >2).Among them,21 miRNAs were up-regulated(miR-106b-3p,miR-584-5p,miR-139-3p,miR-185-3p,miR-625-3p,miR-769-5p,miR-379-5p,miR-23a-3p,miR-425-5p,miR-199a-5p,miR-146b-5p,miR-370-3p,miR-223-3p,miR-382-5p,miR-361-3p,miR-9-3p,miR-589-5p,miR-4665-5p,miR-941,miR-30c-5p,miR-7-5p)and 5 miRNAs were down-regulated(miR-195-5p,miR-29c-3p,miR-15a-5p,miR-4732-5p,miR-34c-5p).5.The results predict the 26 target genes in miTarBase database,miRDB database and Targetscan database that may be regulated by miRNA,and take the intersection of three databases as the final result.We predict that there are 154 target genes that may be regulated by mi RNA.6.The functional analysis of 154 target genes showed that their functions were enriched in 117 go entries and 189 KEGG pathways.Conclusion Stable miRNA can be detected in serum of ASD children.Compared with the normal children,the expression of 26 miRNA in the serum of ASD children was different.The target gene functions regulated by differentially expressed miRNAs are enriched in a variety of biological processes,and the signal transduction pathways involved are closely related to ASD.26 mi RNAs differentially expressed between the two groups can be used as potential biomarkers for early screening of ASD. |
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