Protein Expression Eukaryotic cells W. anomalus

RNAi or RNA interference is a common method to suppress gene expression in vitro/in vivo by utilizing the inherent microRNA machinery, without introducing a total gene knockout. miRNA is the inherent gene silencing machinery which can have more than one mRNA target, whereas siRNA can be designed to target a particular mRNA target. By design, both siRNA and miRNA are 20-25 nucleotides in length. The target sequence for siRNAs is usually located within the open reading frame, between 50 and 100 nucleotides downstream of the start codon. There are two ways in which cells can be transfected with desired RNAi: 1. Direct transfection (with calcium phosphate co-precipitation or cationic lipid-mediated transfection using lipofectamine or oligofectamine), and 2. Making RNAi lentiviral constructs (followed by transformation and transduction). Lentiviral constructs are time-consuming, but provide a more permanent expression of RNAi in the cells and consistent gene silencing. Direct transfection of oligonucleotides provides temporary genetic suppression. Traditional methods like calcium phosphate co-precipitation have challenges like low efficiency, poor reproducibility and cell toxicity. Whereas, cationic lipid-based transfection reagents are able to overcome these challenges, along with applicability to a large variety of eukaryotic cell lines.

Get tips on using Color-coded Prestained Protein Marker, Low Range (1.7-42 kDa) #13070 to perform Protein Ladder Prestained

Get tips on using Color-coded Prestained Protein Marker, High Range (43-315 kDa) #12949 to perform Protein Ladder Prestained

Get tips on using Color-coded Prestained Protein Marker, Broad Range (10-250 kDa) #74124 to perform Protein Ladder Prestained

Get tips on using Prestained Dual Color Protein Molecular Weight Marker (1.7-40 kDa) Molecular Weight Marker to perform Protein Ladder Prestained

Get tips on using heat shock protein family A (Hsp70) member 5 to perform siRNA / miRNA gene silencing Human - PC3 (human prostate cancer cell line) HSPA5 (GRP78)

Get tips on using Anti-TATA binding protein TBP antibody [58C9] - Loading Control (ab61411) to perform Western blotting TBP