Porcine Tissue RNA Extraction Protocols

Seventeen different porcine tissues were collected from three young female siblings. Total RNA was extracted using different protocols depending of the tissue: TRIreagent^®(Molecular Research Centre, inc.) for liver, kidney, thymus, RNeasy lipid kit (Qiagen) for adipose (subcutaneous), cortex cerebri, cerebellum, hippocampus, lymph nodules (jejunal), RNeasy Fibrous Kit (Qiagen) for muscle (longissimus dorsi), heart (muscle), skin (dermis and epidermis) and RNeasy kit (Qiagen) for pancreas, bone marrow, bladder, lung, stomach (mucosal membranes), small intestine (mucosal membranes) according to each manufacturer protocol. Contaminating DNA was degraded by treating each sample with RQ1 RNase-free DNase (Promega) according to the instructions manual, followed by a spin-column purification (Qiagen RNeasy). The total RNA was quantified by optical density and the quality was evaluated by gel electrophoresis. Intact rRNA subunit of 28S and 18S were observed on the gel indicating minimal degradation of the RNA.

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Nygard A.B., Jørgensen C.B., Cirera S, & Fredholm M. (2007). Selection of reference genes for gene expression studies in pig tissues using SYBR green qPCR. BMC Molecular Biology, 8, 67.

Publication 2007

28s rrna Adipose Bladder Bone marrow Cerebellum Cortex cerebri Dermis Dnase Electrophoresis Epidermis Female Fibrous Heart muscle Hippocampus Jejunal Kidney Lipid Liver Lung Lymph nodules Membranes Mucosal Muscle Optical Pancreas Porcine Promega Rna degradation Rnase Siblings Skin Small intestine Stomach Subunit Thymus Tissue

Corresponding Organization : University of Copenhagen

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Protocol cited in 20 other protocols

Variable analysis

independent variables

Tissue type (liver, kidney, thymus, adipose, cortex cerebri, cerebellum, hippocampus, lymph nodules, muscle, heart, skin, pancreas, bone marrow, bladder, lung, stomach, small intestine)

dependent variables

Total RNA quantification by optical density
RNA quality evaluation by gel electrophoresis

control variables

Tissue source (three young female siblings)
RNA extraction protocol (TRIreagent, RNeasy lipid kit, RNeasy Fibrous Kit, RNeasy kit)
DNA degradation (RQ1 RNase-free DNase treatment and spin-column purification)

controls

Positive control: Observation of intact rRNA subunits of 28S and 18S, indicating minimal RNA degradation

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