Site Directed Mutagenesis (SDM) Monkey Deletion COS-7

Get tips on using p21 siRNA (h) to perform siRNA / miRNA gene silencing Human - COV-434 P21

Get tips on using Mouse SDF-1 α / CXCL12 α ELISA Kit to perform ELISA Mouse - SDF-1/CXCL12

Get tips on using Mouse CXCL12/SDF-1 alpha Quantikine ELISA Kit to perform ELISA Mouse - SDF-1/CXCL12

Get tips on using EmbryoMax MEF Conditioned Media to perform Stem cell culture media hESC lines H9, H1

Get tips on using MagNA Pure Compact RNA Isolation Kit to perform RNA isolation / purification Cells - primary human mesenchymal stem cells

Get tips on using ON-TARGETplus Human SMU1 (55234) siRNA - Individual to perform siRNA / miRNA gene silencing Human - HeLa SMU1

Get tips on using Lipofectamine® RNAiMAX Transfection Reagent to perform siRNA / miRNA gene silencing Human - siRNA negative control Lipid

Get tips on using Pierce™ Coomassie (Bradford) Protein Assay Kit to perform Protein quantification Mammalian cells - Human pluripotent stem cells

Get tips on using pcDNA™3.1 (+) Mammalian Expression Vector to perform siRNA / miRNA gene silencing Human - U251 cofilin-1 (CFL1)

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.