Experiment 7: Separation of Proteins byPolyacrylamideGelElectrophoresis(PAGE)口 PAGE: an electrophoretic method that utilizespolyacrylamide gel as the support medium.口It serves as a vital tool for studying macromoleculessuch as proteins and nucleic acids口The gel is a 3D network structure polymerized fromacrylamide (monomers) and methylene bisacrylamide(cross-linker) in the presence of a catalyst.['meerli:n]-CH2
Experiment 7: Separation of Proteins by Polyacrylamide Gel Electrophoresis (PAGE) PAGE: an electrophoretic method that utilizes polyacrylamide gel as the support medium. It serves as a vital tool for studying macromolecules such as proteins and nucleic acids. The gel is a 3D network structure polymerized from acrylamide (monomers) and methylene bisacrylamide (cross-linker) in the presence of a catalyst. ['meθɪliːn] -CH2
l.PolymerizationofAcrylamidePolyacrylamide gel is formed through the polymerization andcrosslinking of the monomer acrylamide (Acr) and the crosslinkerN,N-methylene-bisacrylamide (Bis) under the action of theaccelerator N,N,N,N-tetramethyl ethylenediamine (TEMED) andthe catalyst ammonium persulfate (AP) or riboflavin (vitamin B2)This 3D network structure gel serves as the support medium forelectrophoresis, which is referred to as polyacrylamide gelelectrophoresis (PAGE)
1. Polymerization of Acrylamide Polyacrylamide gel is formed through the polymerization and crosslinking of the monomer acrylamide (Acr) and the crosslinker N,N-methylene-bisacrylamide (Bis) under the action of the accelerator N,N,N,N-tetramethyl ethylenediamine (TEMED) and the catalyst ammonium persulfate (AP) or riboflavin (vitamin B2). This 3D network structure gel serves as the support medium for electrophoresis, which is referred to as polyacrylamide gel electrophoresis (PAGE)
> PhotopolymerizationCatalysed under light excitation (sunlight or daylight)using riboflavin as the catalyst to produce macroporous gel> Chemical polymerizationAP serves as the initiatorTEMED acts as the acceleratorUsed to produce microporous gel
Photopolymerization Catalysed under light excitation (sunlight or daylight), using riboflavin as the catalyst to produce macroporous gel. Chemical polymerization AP serves as the initiator. TEMED acts as the accelerator. Used to produce microporous gel
Reaction:—CH2—CH—[CH2—CH]—CH2O=oCH,=CH—CCONH2NHNHCH2CH2=CH—CONH2+JCH2NHNHCONH2CH,=CHIC=010—CH2—CH—[CH2—CH]x—CH2-BisAcrpolyacrylamide gel
NH CH2=CH—CONH2 + CH2=CH—C CH2=CH—C —CH2—CH—[CH2—CH]x—CH2— —CH2—CH—[CH2—CH]x—CH2— CONH2 C=O CH2 NH NH O O CONH2 C=O NH CH2 Acr Bis polyacrylamide gel Reaction:
2.Principles ofPAGEThree physical effects:> Sample concentration effect> Molecular sieve effect> Charge effect
2. Principles of PAGE Three physical effects: Sample concentration effect Molecular sieve effect Charge effect