6 protocols | |
|---|---|
| Accession | Type |
nucleic acid sequencing protocol | |
Resulting libraries were quality checked on an Agilent DNA 1000 bioanalyzer (Agilent Technologies, South Queensferry, UK) and then clustered onto a paired end flowcell using the Illumina TruSeq Rapid PE Cluster kit at an 8pM concentration. 100 cycle paired-end sequencing was carried out on the Illumina HiSeq 2500 using an Illumina TruSeq Rapid SBS kits (Illumina, Little Chesterford, UK). The Illumina HiSeq 2500 platform generated 13-18 million RNAseq tags per sample (191 million in total), each 100 nucleotides in length, resulting in 46.5 Gb of data. This was carried out at the Edinburgh Genomics facility within Roslin Institute, Midlothian, UK. | |
treatment protocol | |
Cells were cultured with or without LPS for 24 h before RNA was isolated | |
nucleic acid library construction protocol | |
Samples were prepared for mRNA sequencing using 2ug of total RNA starting material following the Illumina TruSeq RNA Sample Preparation v2 kit protocol. | |
nucleic acid extraction protocol | |
Standard Trizol extraction method | |
growth protocol | |
Macrophages and dendritic cells were cultured from bone marrow by standard methods (Garceau et al. 2010 and Wu et al. 2010, respectively), and heterophils isolated from blood by standard methods (Kogut et al. 1995). | |
high throughput sequence alignment protocol | |
Tophat version 2.0.9 was used with Bowtie2 version 2.1.0 for alignment of reads. Reads were aligned to the Galgal 4 genome assembly from the Ensembl 72 release. | |