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Table 1 Pig intestinal organoids

From: Intestinal organoids in farm animals

Segment Application Crypt isolation Growth medium Reference
Duodenum Lipopolysaccharides challenge 30 mM EDTA, 1.5 mM DTT, 4 °C, 20 min + 30 mM EDTA, 10 µM Y27632, warm, 10 min DMEM, N2, B27 without vitamin A, 10 mM HEPES, 2 mM L-glutamine, 1 μg/mL R-spondin1, 100 ng/mL Noggin, 50 ng/mL EGF, 2.5 ng/mL Wnt-3a [60]
  Gene edited pigs 1 mg/mL collagenase, 2 mg/mL BSA, 37 °C, 30 min IntestiCult™ (STEMCELL Technologies, Vancouver, Canada) [37]
Jejunnum Cell lineage identification 30 mM EDTA, 1 mM DTT, 10 µM Y27632, 4 °C, 20 min and 37 °C 10 min DMEM/F12, N-2, B-27 minus vitamin A, Glutamax, 1 mM HEPES, 1 mM NAC, 100 ng/mL rm Noggin, 500 ng/mL rh R-spondin, 50 ng/mL rm EGF, 100 ng/mL rh Wnt3a, 10 µM Y-27632, 10 µM SB202190, 500 nM LY2157299 [39]
  Lentivirus transduction 10 mM EDTA, 1 mM DTT, 4 °C, 30 min DMEM/F-12, 2 mM glutaMAX, 10 mM HEPES, 1 mM NAC, N2, B27, 50 ng/mL rm EGF, 100 ng/mL rm Noggin, 500 ng/mL rh R-spondin 1, Wnt3a CM, 1 mM valproic acid, 10 mM nicotinamide, 2.5 μM CHIR99021, 10 μM Y27632, 10 μM SB202190, 500 nM LY2157299 [25]
  2D monolayers, transcriptomics 2 mM EDTA, 4 °C, 30 min DMEM/F12, 10 mM HEPES, B-27, 1.25 mM NAC, 50 ng/mL rh EGF, 15 nM gastrin, 10 mM nicotinamide, 10 μM p38 MAPK inhibitor, 600 nM A83-01, Noggin CM (15% v/v), R-spondin CM (15% v/v), and Wnt3A CM (30% v/v) [5, 19]
  Mycotoxin challenge 30 mM EDTA, 1.5 mM DTT, 10 µM Y27632, 4 °C, 30 min DMEM, 10% FBS, N2, B27, 1X GlutaMAX, 1 mM NAC, 50 ng/mL EGF, 100 ng/mL Noggin, 500 ng/mL R-spondin1, 10 mM nicotinamide, 10 µM SB202190, 0.5 µM LY2157299, 45% Wnt3a CM, 10 µM Y27632 (2–3 first days) [18]
  Heat Stress 30 mM EDTA, 1.5 mM DTT, 4 °C, 30 min DMEM/F12, Wnt3a CM, rm EGF, rm Noggin, rh R-spondin1, nicotinamide, Y27632, LY2157299, SB202190, CHIR99021, FBS, N2, B27, glutamine, NAC [26]
  Swine enteric virus infection 5 mM EDTA, 4 °C, 1 h IntestiCult™ (STEMCELL Technologies, Vancouver, Canada) [34]
  S. Typhimurium and T. gondii infection 0.8 mM EDTA, 4 °C, 30 min IntestiCult™ (STEMCELL Technologies, Vancouver, Canada) [20]
Ileum Model development Collagenase, 37 °C, 1.5 h 50% (DMEM + 10% FBS), 50% L-WRN CM, 10 µM SB431542, 10 µM Y27632 [14]
  Vitamin A 2 mM EDTA, 4 °C, 30 min Wnt3a, Noggin and R-spondin 1 CM, B27, N2, glutamine, NAC, rm EGF, nicotinamide, SB202190 [45]
  Mucus production 30 mM EDTA, 1 mM DTT, 37 °C, 10–15 min IntestiCult™ (STEMCELL Technologies, Vancouver, Canada) [43]
  Lawsonia intracellularis infection 30 mM EDTA, 1 mM DTT, 37 °C, 5–15 min IntestiCult™ (STEMCELL Technologies, Vancouver, Canada) [36]
Duodenum, Jejunum, Ileum, Colon Porcine epidemic diarrhea virus infection Cell dissociation reagent (STEMCELL Technologies,Vancouver, Canada) IntestiCult™ (STEMCELL Technologies, Vancouver, Canada) [21]
  1. CM: conditioned medium, NAC: N-acetyl cysteine, L-WRN: L cell line engineered to secrete Wnt3a, R-spondin 3 and Noggin, rm: recombinant mouse, rh: recombinant human, FBS: fetal bovine serum, EGF: epidermal growth factor. Antibiotics and antifungals added in the growth medium are not presented.