Puromyocin

Manufactured by Merck Group

Sourced in United States

Puromycin is a laboratory reagent used in cell culture experiments. It functions as a protein synthesis inhibitor, specifically targeting the translation process in eukaryotic cells.

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Lab products found in correlation

Lentiviral particles, by GenePharma (1 mentions) Polybrene, by Merck Group (1 mentions) Lentivirus, by GenePharma (1 mentions) Opti mem media, by Thermo Fisher Scientific (1 mentions) Fugene 6, by Promega (1 mentions) Fetal bovine serum (fbs), by Biowest (1 mentions) Pen strep, by Lonza (1 mentions)

2 protocols using puromyocin

Knockdown of PKR and STAT3 in HepG2 Cells

Cited 1 time

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Short hairpin RNA (shRNA) sequences were obtained from Public TRC Portal (Broad Institute, Cambridge, MA, USA) and lentiviruses expressing the shRNA sequences were synthesized by Shanghai GenePharma Co., Ltd., (Shanghai, China). Polybrene and puromyocin were purchased from Sigma-Aldrich (St. Louis, MO, USA). Transfection of the HepG2 cells with lentiviral particles (Shanghai GenePharma Co., Ltd.)was conducted as described previously (23 (link)). Titration of the lentiviruses was performed (not shown), and the lowest functional quantities of the virus (MOI=5) and Polybrene were subsequently applied in the LOF experiments. The shRNA target sequences for PKR were as follows: shPKR-1, 5′-GCTGAACTTCTTCATGTATGT-3′ and shPKR-2, 5′-GAGGCGAGAAACTAGACAAAG-3′. The shRNA target sequence for STAT3 was 5′-GCACAATCTACGAAGAATCAA-3′.

WANG X., DONG J.H., ZHANG W.Z., LENG J.J., CAI S.W., CHEN M.Y, & YANG X. (2014). Double stranded RNA-dependent protein kinase promotes the tumorigenic phenotype in HepG2 hepatocellular carcinoma cells by activating STAT3. Oncology Letters, 8(6), 2762-2768.

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Establishing GFP-expressing HS-5 Stromal Cells

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Human bone marrow stromal cell line, HS-5 cells (ATCC) were cultured in DMEM (Dulbecco's modified Eagle's medium, ATCC) supplemented with 10% fetal bovine serum (FBS) (Biowest) and 1% PenStrep (Lonza). The cells were cultured in T-75 flasks in 5% CO₂ atmosphere at 37°C and split in a 1:5 ratio every 3-4 days. For confocal fluorescent imaging HS-5 cells were transfected with Green Fluorescent Protein (GFP)²⁴. Briefly, 24 hours prior to transfection, HS-5 cells were seeded and cultured in 6 well plates. Once cell confluency reached 60% a transfection reagent Fugene 6 (Promega), was first mixed with serum-free Opti-Mem media (Invitrogen) and sequentially with DNA (pLenti-GIII-GFP) (Applied Biological Materials) according to the manufacturer's instructions. This mixture was carefully added dropwise and then incubated at 37°C for 24 – 48 hours. After verifying HS-5 was successfully transfected with GFP, transfected cells were selected and cultured in media supplemented with puromyocin (1μg/ml) (Sigma-Aldrich).

Kim J., Kong Y.P., Niedzielski S.M., Singh R.K., Putnam A.J, & Shikanov A. (2016). Characterization of the crosslinking kinetics of multi-arm poly(ethylene glycol) hydrogels formed via Michael-type addition. Soft matter, 12(7), 2076-2085.

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