Computational protocol: Transcriptional analysis of liver from chickens with fast (meat bird), moderate (F1 layer x meat bird cross) and low (layer bird) growth potential

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Protocol publication

[…] Reads were returned in fastq format. FastQC and adaptor sequences were trimmed from the 3′ end of reads with Cutadapt []. Hisat2 [] was used to map reads to the reference genome Galgal5.0 (ftp://ftp.ncbi.nlm.nih.gov/genomes/Gallus_gallus). Duplicate reads were then removed. Stringtie [] was used to define the transcripts from the read mappings for each sample, and to merge the transcript definitions for all samples. Transcripts were cleaned up using in-house scripts. The number of raw read counts were calculated for each transcript and sample using the function feature Counts of the R package Rsubread []. Another R package, edgeR [] was used to analyse differential gene expression using normalised counts per million transcripts (CPM) to correct for varying depth of sequence among samples. Transcript data were aggregated by gene. Genes where the maximum CPM was < 1 were removed. Gross transcriptome relationships between the three types of bird were analysed by multidimensional scaling of the CPMs. [...] Functional enrichment of the DE genes between meat bird vs layer, meat bird vs cross and layer vs cross and DE between all three groups was conducted for gene ontology (GO) terms and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways using the web based tools in DAVID [, ]. Only GO terms and KEGG pathways with P < 0.05 were taken into account as significantly enriched among the DE genes. Phenotypic data, including bodyweight, bodyweight gain and liver weights (normalised and actual), were analysed by a one-way ANOVA using SPSS (IBM SPSS Statistics 22). Gene expression levels were correlated with individual bodyweights (all three groups combined) using a Pearson’s correlation in SPSS (IBM SPSS Statistics 22). […]

Pipeline specifications

Software tools FastQC, cutadapt, HISAT2, StringTie, Subread, edgeR, DAVID, SPSS
Databases KEGG
Applications Miscellaneous, RNA-seq analysis
Organisms Gallus gallus, Homo sapiens
Chemicals Glucose