dna-isolation-purification-bacteria-gram-positive-clostridium-difficile

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Plasmid Isolation Clostridium difficile Experiment

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.

DNA Plasmid Isolation Clostridium difficile

RNA isolation / purification Bacteria - Gram positive Streptococcus pneumoniae Experiment

The biggest problem in isolating RNA from gram-positive bacteria is the disruption of the cell wall. A lot of protocols employ enzymatic digestion (pretreatment) which may affect gene expression patterns of certain genes. Therefore physical disruption using beads could be the best alternative.

RNA RNA isolation / purification Bacteria Gram positive Streptococcus pneumoniae

RNA isolation / purification Bacteria - Gram positive Mycobacterium tuberculosis Experiment

RNA RNA isolation / purification Bacteria Gram positive Mycobacterium tuberculosis

RNA isolation / purification Bacteria - Gram positive Enterococcus faecalis Experiment

RNA RNA isolation / purification Bacteria Gram positive Enterococcus faecalis

RNA isolation / purification Bacteria - Gram positive Bacillus anthracis Experiment

RNA RNA isolation / purification Bacteria Gram positive Bacillus anthracis

RNA isolation / purification Bacteria - Gram positive Lactobacillus amylovorus Experiment

RNA RNA isolation / purification Bacteria Gram positive Lactobacillus amylovorus

RNA isolation / purification Bacteria - Gram positive Listeria monocytogens Experiment

RNA RNA isolation / purification Bacteria Gram positive Listeria monocytogens

RNA isolation / purification Bacteria - Gram positive Staphylococcus epidermidis Experiment

RNA RNA isolation / purification Bacteria Gram positive Staphylococcus epidermidis

RNA isolation / purification Bacteria - Gram positive Staphylococcus saprophycitius Experiment

RNA RNA isolation / purification Bacteria Gram positive Staphylococcus saprophycitius

RNA isolation / purification Bacteria - Gram positive Streptococcus pyogenes Experiment

RNA RNA isolation / purification Bacteria Gram positive Streptococcus pyogenes

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