Maxiprep FAQ

Maxiprep FAQ

• 
  Q: What does TENS buffer do?

  • Contains Tris-HCl, EDTA, and NaOH, and SDS

  • Tris-HCl is the buffer part, maintains a stable pH to protect the DNA

  • EDTA inhibits enzymes that may break down DNA

  • Na+ neutralizes negatively charge DNA, makes it less soluble in water but more soluble in ethanol, which is used later

  • Overall it is a lysis buffer that breaks open the bacterial cells to expose the DNA we want

• Q: What does sodium acetate buffer do?

  • Another salt to neutralize charges on the DNA molecule, making it less hydrophilic

• Q: What is RNase?

  • A ribonuclease

  • Degrades single stranded RNA, makes sure only DNA is present in sample

• Q: What does PCI do?

  • PCI is an organic mixture and allows the partitioning of cell debris and lipids into an organic layer, while DNA remains in the aqueous layer

• Q: What does chloroform do?

  • Helps denature proteins as well as aids in partitioning to separate DNA into the aqueous layer while undesired cell products are in the organic layer

• Q: What is the purpose of ethanol?

  • Addition of ethanol removes the salts that were previously added to the DNA

  • DNA without the salts is not soluble in ethanol, so it is precipitated out of the solution

  • The ethanol is then decanted and allowed to evaporate, leaving purified DNA in the tube :)