Protein expression and purification Insect cells Sf9

Get tips on using CelLytic™ M to perform Protein isolation Mammalian cells - SH-SY5Y

Get tips on using CelLytic™ M to perform Protein isolation Mammalian cells - BHK-21

Get tips on using Gentra Puregene Buccal Cell Kit (100) to perform DNA isolation / purification Cells - Primary cells Buccal cells

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 Gentra Puregene Blood Kit to perform DNA isolation / purification Cells - Primary cells Bone marrow mononuclear cells

Get tips on using TRIzol Reagent to perform RNA isolation / purification Cells - primary human mononuclear cells

Get tips on using TRIzol Reagent to perform RNA isolation / purification Cells - primary human endothelial cells