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Exploration and analysis of high-throughput sequence data of bioinformatics: analysis of transcriptome studies and Schistosoma japonicum Hox gene family in zebrafish embryonic development

Author: GuJianLei
Tutor: ZhouYan
School: Fudan University
Course: Microbiology
Keywords: Zebrafish Transtriptome Deep-seq Embryogenesis Hox genes Schistosomajaponicum
CLC: Q811.4
Type: Master's thesis
Year: 2011
Downloads: 99
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Abstract


Zebrafish is a convenient research model species for development biology, because of their rapid development progresses. It was difficult to study the transcriptome of zebrafish due to the limitations of microarray profiling technologies. However, the deep-sequencing technologies offer several advantages over transcriptome analysis by microarray including increased throughput and resolution. Based on SOLiD3system and series bioinformatics analysis, we launched a dynamic transcriptome analysis among9stages of zebrafish embryogenesis.We generated10libraries from9stages and sequenced them to a depth of almost3billion reads. And we mapped the reads to the zebrafish unigene library (uniq library, build117). We used custom mapping strategy to maximize reads-mapping coverage; finally, more than34.82%of total reads were matched in the sense orientation. Our transcriptome profile included89.2%unigenes with high-level mapping to reference library. We calculated the relative expression level by counting the number of reads mapping to each unigene, and normalized raw data via RPKM method. We used reads counts and statistic test method to define significant differential expression between adjacent libararies. We observed39,824significant (p-value<=0.001) differentially expressed genes in9stages. Furthermore, in order to place the uncharacterized profile into biological context, we clustered the reference sequences into functional groups via several databases (including Gene ontology, pfam and KEGG pathway). Enrichment analysis revealed that a large proportion of genes are not expressed in the early embryogenesis period. For example, in the cleavage and blastula periods, the cell cycle, transcription, DNA/protein modification and gene-silence-related genes exhibit high expression levels. However, a large number of genes were beginning to be expressed since the gastrula period. The clustering analysis shows the dynamic variance during embryo development. For example, the development of somite and organ were started at the early somite period, and since the60hpf stage, Tissues (especially the neuro system) most likely do their specific functions.In Chapter2, we took a genome-wide investigation for Hox genes in Schistosoma japonicum. Hox genes are characterized by a highly conserved peptide domain and contribute to the antero-posterior axis patterning during embryogenesis. Due to their central role in development biology, these genes have been widely studied in various animals. Based on the published genome assembly and re-sequencing project of Schistosoma japonicum, we present genome-wide characterization and comparative genomic analysis of the Hox gene family within this parasitic flatworm. Eight Hox genes were identified and validated in our investigation. Comparative analysis revealed that these genes were distributed among seven orthology groups of the Hox gene family, which were characterized as:Hoxl, Hox2, Hox3, Hox4, Lox5, Lox4and Post-2. Semi-quantitative RT-PCR was used to investigate the expression pattern of four Hox genes in the egg, miracidias, schistosomulae and adult stages. Our study is the first genome-wide characterization of Hox genes in S. japonicum, providing useful information about this important gene family in parasitic flatworms.

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