title: 3609
organism: Sus scrofa
description: Protocols: All pigs were from parity 2 of generation 9 of the two lines divergently selected for residual feed intake: the low and high RFI lines. Blood samples were collected from the jugular vein into TempusTM Blood RNA tubes (Life Technologies, Grand Island, NY) for long-term storage at -80 C from 233 post-weaning piglets of the two lines in this age range. Meanwhile, blood from those animals was also collected into EDTA tubes (BD, Franklin Lakes, NJ) and kept at 4 C before CBC tests, which were performed on the bleeding day, if possible, or the next morning. At 107.0 8.3 (mean standard deviation) days of age and 42.2 7.2 kg of body weight (BW), 88 gilts and 78 barrows were randomly assigned to 12 mixed-line, mixed-sex finishing pens with electronic single-space feeders (FIRE, Osborne Industries Inc., Osborne, KS) for feed efficiency test, with 6 pens being randomly assigned to either of two diets: a high-fiber, low-energy diet (HFD) and a low-fiber, high-energy diet (LFD). The animals on feed efficiency test consisted of 21 barrows and 22 gilts from the low RFI line plus 14 barrows and 23 gilts from the high RFI line fed the HFD, and 23 barrows and 23 gilts from the low RFI line plus 20 barrows and 20 gilts from the high RFI line fed the LFD. All pigs had ad libitum access to feed and water. Individual feed intake was real-time recorded, body weight was recorded biweekly, and backfat (BF) depth above the 10th rib and loin muscle area were recorded at the end of the test, when the pigs were 227.0 1.4 days of age and 127.7 8.8 kg of body weight. RFI of individual pigs were calculated. In detail, for the low RFI line, we selected eight samples from animals with extremely low RFI values on the LFD and eight samples from animals with extremely low RFI values on the HFD (designated as low RFI group); while for the high RFI line, we selected eight samples from animals with extremely high RFI values on the LFD and eight samples from animals with extremely high RFI values on the HFD (designated as high RFI group) (Figure 1). Total RNA extraction of the 32 samples was processed in four batches such that for each batch, two samples were randomly selected from each line by diet combination. Within each batch, the processing order of samples was randomized beforehand and followed in all subsequent procedures where blocking was necessary. The total RNA was extracted from blood samples using preserved blood RNA purification kit I (Norgen Biotek Corp, Thorold, Ontario) by following the kits manual. On-column DNA digestion was performed as described using DNase I (Qiagen, Valencia, CA). Globin transcripts (HBB [ENSSSCG00000007978] and HBA [ENSSSCG00000014725]) were depleted by following an RNase H-mediated method [40]. The quantity and integrity of the RNA were determined by using Nanodrop 2000 (Thermo Scientific, Waltham, MA) and Bioanalyzer 2100 (Agilent Technologies, Santa Clara, CA) before and after globin depletion. Library construction and sequencing were performed by the DNA facility at Iowa State University. Briefly, the RNA-seq libraries were constructed using the Illumina TruSeq RNA Sample Preparation Kit v2 (Illumina, San Diego, CA) according to manufacturers instructions. For each sample, poly (A)-containing transcripts was enriched with oligo-dT-coated magnetic beads from 0.7 to 2 g of total RNA with globin transcript depleted. The enriched RNA was fragmented by heat and reverse transcribed with hexamer random primers. For each sample, adapters with unique barcodes were ligated to the end-polished cDNA fragments. The libraries were linearly amplified by PCR, size selected and quantitated. The individual libraries were diluted to 2 nM and pooled in approximately equimolar amounts according to the processing batches mentioned above with 8 libraries per pool, except for one pool of 7 libraries.
attributes.tag: ENA first public
attributes.value: 2016-01-15
attributes.tag: ENA last update
attributes.value: 2016-01-05
attributes.tag: External Id
attributes.value: SAMEA3720871
attributes.tag: INSDC center alias
attributes.value: Iowa State University
attributes.tag: INSDC center name
attributes.value: Iowa State University
attributes.tag: INSDC first public
attributes.value: 2016-01-15T17:02:21Z
attributes.tag: INSDC last update
attributes.value: 2016-01-05T16:25:52Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-4179:3609
attributes.tag: age
attributes.value: 5-6
attributes.tag: basophil
attributes.value: 80
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: cell type
attributes.value: whole blood
attributes.tag: common name
attributes.value: pig
attributes.tag: developmental stage
attributes.value: postweaning
attributes.tag: diet fed
attributes.value: low-fiber, high energy diet
attributes.tag: eosinophil
attributes.value: 430
attributes.tag: growth condition
attributes.value: ad libitum
attributes.tag: lymphocyte
attributes.value: 11050
attributes.tag: monocyte
attributes.value: 1440
attributes.tag: neutrophil
attributes.value: 7200
attributes.tag: phenotype
attributes.value: low RFI
attributes.tag: rfi value
attributes.value: -0.957876492
attributes.tag: rna concentration after globin depletion
attributes.value: 215
attributes.tag: rna concentration before globin depletion
attributes.value: 945.81
attributes.tag: rna processing batch
attributes.value: 2
attributes.tag: rna rin values after globin depletion
attributes.value: 7.8
attributes.tag: rna rin values before globin depletion
attributes.value: 8.9
attributes.tag: sample name
attributes.value: E-MTAB-4179:3609
attributes.tag: sex
attributes.value: female