siRNA / miRNA gene silencing Human hES cell line H1 (WA01)

Get tips on using ROS-Glo™ H2O2 Assay to perform ROS assay cell type - Huh7

Get tips on using SQSTM1/p62 (D5E2) Rabbit mAb to perform Autophagy assay cell type - A2780

Get tips on using SQSTM1/p62 (D1Q5S) Rabbit mAb to perform Autophagy assay cell type - A549

Get tips on using LC3 Antibody (APG8B) (N-term) to perform Autophagy assay cell type - A549

Get tips on using Annexin V-FITC Apoptosis Kit to perform Apoptosis assay cell type - K562

Get tips on using Autophagy/Cytotoxicity Dual Staining Kit to perform Autophagy assay cell type - Macrophages

Get tips on using Gibco™DMEM/F-12, no glutamine to perform Stem cell Differentiation media hPSCs or iPSCs differentiation into Lung progenitor cells

DNA-protein interactions are studied by using ChIP. The basic steps in this technique are crosslinking, sonication, immunoprecipitation, and analysis of the immunoprecipitated DNA. During ChIP, if chromatin is under-fragmented or fragments are too large which can lead to the increased background and lower resolution. Shorter cross-linking times (5-10 min) and/or lower formaldehyde concentrations (<1%) may improve shearing efficiency. If Chromatin is over-fragmented, then optimize shearing conditions for each cell type to improve ChIP efficiency. Over-sonication of chromatin may disrupt chromatin integrity and denature antibody epitopes. If you do not see any product or very little product in the input PCR reactions, add 5–10 μg chromatin per IP.

The process of RNA extraction from bacteria, in general, involves an RNA-protective, effective lysis of bacterial cell wall (which may pose difficulties). EDTA promotes loss of outer membrane to provide lysozyme with access to peptidoglycan. Another common method for cell wall lysis is mechanical disruption using a homogenizer (applied for gram-positive bacteria and some strains of gram-negative bacteria). Following lysis, it is necessary to disrupt protein-nucleic acid interactions, which can be achieved by adding sodium dodecyl sulfate (SDS). Next step involves using phenol-chloroform-isoamyl alcohol extraction, where RNA can be obtained from the bottom organic phase, the top phase consists of DNA and the interphase contains proteins. Isoamyl alcohol is an inert and optional addition to this mixture and is added as an anti-foaming reagent to reduce the interphase. Following RNA extraction, the samples should be checked for its quality by gel electrophoresis (23S and 16S rRNAs and 5s rRNA and tRNA bands) or UV spectrophotometric or fluorescence methods.

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