DNA isolation / purification

Get tips on using PowerSoil® DNA isolation - DNeasy PowerSoil Pro Kit to perform DNA isolation / purification Bacteria - Gram positive Lactobacillus amylovorus

Get tips on using Wizard® Plus Maxipreps DNA Purification System to perform Plasmid Isolation E. coli DH5α

Get tips on using DNA Isolation Kit for Cells and Tissues to perform DNA isolation / purification Cells - Primary cells Human primary keratinocytes

Get tips on using Wizard® Plus Midipreps DNA Purification System Technical Bulletin to perform Plasmid Isolation Proteus mirabilis

Get tips on using Wizard® Plus SV Minipreps DNA Purification System Technical Bulletin to perform Plasmid Isolation Streptomyces spp

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 Apoptotic DNA Ladder Isolation Kit to perform DNA Damage Assay U266

Get tips on using Apoptotic DNA Ladder Isolation Kit to perform DNA Damage Assay U-87 MG

Get tips on using PowerSoil® DNA isolation to perform AAA for reviews

When extracting nucleic acids from cell cultures, thorough homogenization of cells via vortexing in lysis buffer is very necessary. Choose the best RNA isolation method keeping in mind the downstream applications, generally, column-based isolations result in clean and concentrated RNA samples. Downstream applications like sequencing and cDNA synthesis require high-quality RNA, always treat the samples with DNases and check their integrity by running a gel.