siRNA / RNAi /miRNA transfection Human Cells HeLa

Get tips on using RNeasy Mini Kit to perform RNA isolation / purification Cells - primary human chondrocytes

Get tips on using RNeasy Mini Kit to perform RNA isolation / purification Cells - primary human keratinocytes

Get tips on using ROS-ID® Total ROS/Superoxide detection kit to perform ROS assay cell type - A549 human adenocarcinomic human alveolar basal epithelial cells

Get tips on using Live-Dead Cell Staining Kit (BioVision) to perform Live / Dead assay mammalian cells - MDA-MB-231 human breast cancer cells

Get tips on using LIVE/DEAD™ Cell Imaging Kit to perform Live / Dead assay mammalian cells - MDA-MB-231 human breast cancer cells

Get tips on using Purified Mouse Anti-Human ZO-1 Clone 1/ZO-1 (RUO) to perform Western blotting ZO-1

Get tips on using Cytoselect™ Cell Viability and Cytotoxicity Assay to perform Live / Dead assay mammalian cells - MCF-7 human breast cancer cells

Get tips on using RIPA Buffer to perform Protein isolation Mammalian cells - Human lung fibroblasts

The RNA-guided CRISPR-Cas9 nuclease system has revolutionized the genome editing practices. For the most part, the Cas9-mediated genome editing is performed either via nonhomologous end joining (NHEJ) or homology-directed repair (HDR) in mammalian cells, However, designing of specific sgRNAs and minimizing off-target cleavage mediated mutagenesis are the major challenges in CRISPR-Cas based genome editing. To circumvent these issues, we can take advantages of many available tools and approaches for sgRNA construction and delivery.

Protein isolation is a technique that involves isolation and/ or purification of protein from cells or tissues via chromatography or electrophoresis. The major challenges in protein isolation include: 1. The concentration of proteins in cells is variable and tends to be small for some intracellular proteins. Unlike nucleic acids, proteins cannot be amplified. 2. Proteins are more unstable than nucleic acids. They are easily denatured under suboptimal temperature, pH or salt concentrations. 3. Finally, no generalized technique/protocol can be applied for protein isolation. Proteins may have different electrostatic (number of positively or negatively charged amino acids) or hydrophobic properties. Therefore, protein purification requires multiple steps depending on their charge (a negatively charged resin/column for positively charged proteins and vice-versa), dissolution (using detergents) and unlike in the case of DNA and RNA, instead of using salts, proteins should be isolated by isoelectric precipitation.