Total RNA was isolated from in vivo porcine fetal lungs on day 37 of gestation using the mirVanaTM miRNA isolation kit (Ambion, Grand Island, NY USA). Total RNA was tested on an Agilent Model 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA USA). Samples with an RNA integrity number (RIN) greater than 8.0 were selected for further processing.
Libraries were prepared using the TruSeq RNA Sample Prep (Illumina, San Diego, CA) and submitted to the University of Iowa DNA Facility for deep sequencing. 12 paired-end DNA libraries were sequenced to an average depth of 212 million read pairs (range of 179–241 million) with 100 base reads. The sequences were aligned to the Sus scrofa genome (release 10.2) using TopHat (v2.0.10) and known genes were annotated using Ensembl (release 74). Gene expression differences between CF (5 replicates) and non-CF (7 replicates) groups were analyzed using Cuffdiff (v2.1.1).(21 (link))
Libraries were prepared using the TruSeq RNA Sample Prep (Illumina, San Diego, CA) and submitted to the University of Iowa DNA Facility for deep sequencing. 12 paired-end DNA libraries were sequenced to an average depth of 212 million read pairs (range of 179–241 million) with 100 base reads. The sequences were aligned to the Sus scrofa genome (release 10.2) using TopHat (v2.0.10) and known genes were annotated using Ensembl (release 74). Gene expression differences between CF (5 replicates) and non-CF (7 replicates) groups were analyzed using Cuffdiff (v2.1.1).(21 (link))
