GSE60497
Quantitative Analysis of differentially expressed genes between HH10-HH13 stages of chick embryo
Purpose: The aim of this study is to analyse and compare the differentially expressed genes between the developmental stages and to construct heat maps with the data generated. Methods: The trancriptome sequencing was performed using Illumina HiSeq 2000 ® platform. The sequence reads were aligned to the reference genome of chicken (Galgal4) downloaded from Ensembl Release 75 database. Alignment was performed using TopHat (v2.0.8) program to the Ensembl Release 75 gene model of chicken with default parameter settings. After alignment, Cuffdiff (v2.2.0) program was used for performing differential expression analysis. Pathway analysis of differentially expressed genes was performed using DAVID program. Results: Comparison of HH11 vs HH10 showed that gas transport, vascular and blood vessel development, extracellular matrix organization and tube morphogenesis biological processes (BPs) are differentially expressed. Among these, the up-regulated genes are related to extracellular matrix whereas gas transport, vascular and blood vessel development and others are related to down-regulated. Comparison of HH12 vs HH11 revealed that gas transport, iron ion transport and PPAR signaling pathway are enriched for differentially expressed genes. Only up-regulated genes show enrichment of these processes and pathway. It is interesting to note that while gas transport group genes are down-regulated from HH10 stage to HH11 stage, this group of genes are up-regulated while moving from HH11 to HH12. Comparison of HH13 vs HH12 indicated a large number of differentially expressed biological processes and pathways including blood vessel development, vasculature development, cell adhesion, heart development and morphogenesis, extracellular matrix organization, morphogenesis of a branching structure, PPAR signaling pathway, vascular smooth muscle contraction. Overall, we observe a large number of changes occurring between HH12 and HH13 stage. Conclusions: Our study represents the first detailed analysis of differential expression of genes within a stipulated period of time (30-45 hours of egg incubation). Switching on and off of genes happenieng during early vasculogensis is clearly generated by RNA-sequencing. The data analysis reported here will serve as a reference for future investigations on the expression of genes in various other stages . Our results from NGS were in accordance with our experimental data, strongly validating the expression profile. We conclude that genome wide RNA-seq explored the differential expression of genes and pathway interlinked between the stages which permitted us to understand the early vasculogenesis process. Overall design: Analyzing the mRNA profiles of HH10-13 stages of chick embryo were generated by deep sequencing, using Illumina HiSeq 2000 platform.