Computational protocol: Prosteatotic and Protective Components in a Unique Model of Fatty Liver: Gut Microbiota and Suppressed Complement System

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Protocol publication

[…] The mRNA sequencing was performed as previously described with minor modifications. Briefly, RNA samples were prepared using Illumina TrueSeq RNA Sample Preparation Kit (Illumina Inc., San Diego, USA). The concentrations of liver RNA samples from different individuals were adjusted to 10 nM. Sequencing of the cDNA was performed by Personalbio (Shanghai, China) using the Illumina Hiseq system.For mRNA analysis, the raw reads obtained from sequencing were filtered using Fast QC software. The clean reads were then used for the de novo assembly of transcripts by Trinity software. The obtained transcripts were mapped to the chicken (Gallus gallus, Galgal4 version) and duck (Anser platypus, BGI_duck_1.0 version) genomes retrieved from the following website: https://www.ensembl.org using NCBI Blast (NCBI Blast + v2.2.26), respectively. The mapping to the duck genome was a supplement to that to the chicken genome. The mapped reads of each gene were calculated and presented as Reads Per Kb per Million reads (RPKM). The significance was determined using DESeq (http://bioconductor.org/packages/release/bioc/html/DESeq.html). Genes with max RPKM value >2, fold change of treatment over control >2 or < 0.5, and P-value < 0.05 were assigned as differential expressed genes (DEGs). Kyoto Encyclopedia of Genes and Genomes (KEGG) Automatic Annotation Server (KAAS, http://www.genome.jp/tools/kaas/) was used to perform signaling pathway analysis. […]

Pipeline specifications

Software tools Trinity, BLASTN, DESeq, KAAS
Application RNA-seq analysis
Diseases Fatty Liver, Immune System Diseases, Wounds and Injuries
Chemicals Lactic Acid