Next Generation Sequencing Facilitates Quantitative Analysis of A549 Control and A549 infected with Zika virus Transcriptomes. Next Generation Sequencing Facilitates Quantitative Analysis of A549 Control and A549 infected with Zika virus Transcriptomes

Purpose:The goals of this study to compare NGS-derived transcriptome profiling (RNA-seq) to examine differentially expressed genes berween A549 infected with Zika virus and A549 Control . Methods: A549 cells mRNA profiles of A549 cells-infected with Zika virus and aun-infected A549 cells were generated by deep sequencing, in triplicate, using Illumina Hiseq4000. The mapped reads of each sample were assembled by StringTie (v1.3.1) (Mihaela Pertea.et al. 2016) in a reference-based approach. StringTie uses a novel network flow algorithm as well as an optional de novo assembly step to assemble and quantitate full-length transcripts representing multiple splice variants for each gene locus. Results: Using an optimized data analysis workflow, we mapped about identified 11124 transcripts in the A549 cells of control and Zika virus with Cuffcompare workflow. RNA-seq data confirmed stable expression of 20 known housekeeping genes, and 10 of these were validated with qRT–PCR. Approximately 10% of the transcripts showed differential expression between the control and DV, with a fold change >1 and p value <0.05. Altered expression of 25 genes was confirmed with qRT–PCR, demonstrating the high degree of sensitivity of the RNA-seq method. Conclusions: Our study represents the first detailed analysis of A549 cells transcriptomes, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA content within a cell. We conclude that RNA-seq based transcriptome characterization would expedite genetic network analyses and permit the dissection of complex biologic functions. Overall design: A549 cells mRNA profiles of un-infected A549 cells (C) and Zika virus-infected A549 cells (ZV) were generated by deep sequencing, in triplicate, using Illumina HISEQ4000.