Question

Explain the role/function of each of the chemicals found in the three main solutions of the traditional alkaline lysis protocol (GTE, SDS/NaOH, and KOAc).

Answer 1

GTE represents; Glucose, Tris hydroxymethyl aminomethane, ethylenediaminetetraacetic acid

SDS/NaOH represents; sodium dodecyl sulphate and sodium hydroxide.

KOAc represents; Potassium acetate

Step-by-step explanation:

The GTE is used for the resuspension of bacterial cell after their growth and before lyses. Stable environment is provided to cell by GTE solution such as glucose helps in maintaining the osmolarity (similar to inside of cell), tris is involved in maintaining the pH and EDTA chelates the metal ions and prevent them from damaging the plasmid and cell wall.

SDS and NaOH are used for the lyses of the after the resuspension. The plasma membrane is destabilized by SDS. NaOH is involved in the breaking of the cell wall as the hydrogen bonds. It breaks the hydrogen bonds present between double stranded DNA and convert to single stranded DNA. That is why it is called as alkaline lysis.

KOAc is used to decreased the alkalinity of the solution created by NaOH so that hydrogen bonds between single stranded DNA can reestablish and it can form double stranded DNA.