CRISPR Mouse Deletion RMA cells

Get tips on using KAPA RNA HyperPrep Kit with RiboErase (HMR) to perform RNA sequencing Mouse - ESCs (Embryonic Stem Cells)

Get tips on using Quant-iT™ RiboGreen™ RNA Assay Kit to perform RNA quantification Fuorimetric - mouse blood cells

Get tips on using Quant-iT™ RiboGreen™ RNA Assay Kit to perform RNA quantification Fuorimetric - mouse glial cells

Get tips on using T-PER™ Tissue Protein Extraction Reagent to perform Protein isolation Mammalian cells - Mouse Epididymal fat

Get tips on using Luminescent β-galactosidase Detection Kit II to perform Reporter gene assay β-galactosidase substrates - mouse mesenchymal stem cells

Human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) have been greatly used for studies on embryonic development and cell differentiation.iPSCs provide a stable source for either self-renewal or differentiation into suitable cells when cultured in a particular environment. Pluripotent cell culture was originally started by deriving cells from inner cell mass (ICM) from pre-implanted blastocysts, these were called embryonic stem cells. These cells after isolation can be grown on traditional extracellular matrices (like mouse embryonic fibroblasts, MEFs) or feeder-free culture systems. DMEM/F12 has been the most commonly used basal media in the culture of pluripotent cells. These cells are cultured at normal atmospheric oxygen levels, 21%, however, some studies have proposed that 4% oxygen tension may be better for hESC growth. Higher D-glucose concentration (4.2g/l) and osmolarity (320mOsm) that mimics the natural environment of embryonic tissue are optimal for the growth of hESCs. Supplements like N2 and/or B-27, in the presence of growth factors like bFGF, have been shown to increase pluripotency of these cells. bFGF, FGF2 and other ligands of receptor tyrosine kinases like IGF are also required or maintain self-renewal ability of these cells. TGF𝛃1, by its activation of SMAD2/3 signalling, also represses differentiation of iPSCs. Other compounds like ROCK inhibitors reduce blebbing and apoptosis in these cells to maintain their clonogenicity. However, an inhibitor for LIF (leukaemia inhibitory factor, which is one of the pluripotent genes) has an opposing effect. Therefore, it is important to understand the culture conditions and media composition that affect downstream signalling in hESCs or iPSCs that may lead to their differentiation.

Isolating DNA from tissues and paraffin-embedded tissue samples can be challenging as double-stranded DNA is physically fragile and highly susceptible to exo- and endonucleases. The best solution is to slice the tissues into smaller pieces and make a homogenate solution (using tissue homogenizer or grinding liquid nitrogen frozen samples) in the presence of DNAse inhibitors. Further, extracting DNA from the nucleus need specific methods by combining physical, mechanical and chemical lysis approaches,

Get tips on using Live/Dead cell Staining Kit II to perform Live / Dead assay mammalian cells - mouse iPSC

Get tips on using Live and Dead Cell Assay (Abcam) to perform Live / Dead assay mammalian cells - mouse keratinocytes

Get tips on using DMEM/F-12 to perform 3D Cell Culture Media Mouse primary breast ephitelial cells-Mammospheres