siRNA / miRNA gene silencing Human Primary Endometrial Stromal Cells IGFBP1 (Insuline-like growth factor binding protein-1)

Plasmid isolation is an important technique in molecular biology or any kind of genetic editing. It involves amplifying plasmids overnight by transforming them into competent bacterial cells. The desired colonies of these bacteria can then be grown in shaker cultures, at appropriate shaking speed, oxygen availability and temperature. These liquid cultures can then be ultracentrifuged to pellet the bacteria, which are then used for plasmid isolation. The bacteria are first resuspended in a buffer, then lysed, neutralized, purified in a column, eluted, precipitated with ethanol and then resuspended. During plasmid isolation, it is important to lyse cells quickly because lysing bacteria for too long may lead to irreversible denaturing of the plasmid. Usually, alkaline lysis is used for isolation because it is a mild treatment. It isolates plasmid DNA and other cell components such as proteins by breaking cells apart with an alkaline solution. Precipitation removes the proteins, and the plasmid DNA recovers with alcohol precipitation. Resuspension and lysis buffers should be mixed thoroughly in order to prevent the DNA from breaking into smaller fragments. This is because broken gDNA can reanneal and remain in the solution, without binding to the column.

Get tips on using GeneJuice® Transfection Reagent to perform DNA transfection Mammalian cells - Primary cells Human osteoblasts

Get tips on using EGMTM -2 MV Microvascular Endothelial Cell Growth Medium-2 BulletKitTM to perform Stem cell culture media hPericytes

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

Get tips on using JetPrime to perform DNA transfection Mammalian cells - Primary cells Human chondrocytes