Study on SDS Polyacrylamide Gel Electrophoresis for the Analysis of Robinia pseudoacacia L.. Callus

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  • Chen Ren
    Third Course of Forestry Department, Faculty of Agriculture, Kyushu University
  • Gyokusen Koichiro
    Third Course of Forestry Department, Faculty of Agriculture, Kyushu University
  • Saito Akira
    Third Course of Forestry Department, Faculty of Agriculture, Kyushu University

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Other Title
  • ニセアカシア(Robinia pseudoacacia L.)のカルス分析用SDSゲル電気泳動法の検討
  • ニセアカシア Robinia pseudoacacia L.ノ カルス ブンセ

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Description

The method of SDS (sodium dodecyl sulfate) polyacrylamide gel electrophoresis for the analysis of Robinia pseudoacacia L. callus was established. To protect proteins in the sample from being proteolyzed, denatured or modified, and to dissolve insoluble proteins, addition of DTT (dithiothreitol) and urea to the final concentration of 0.375mM and 8M respectively was necessary for the sample preparation. Addition of 0.025% of ammonium peroxodisulfate and TEMED (N, N, N', N' -tetramethylethylenediamine) to the separating gel solution and filling the gel at the impregnation speed of 8mm/min. at room temperature was appropriate for the gradient (7.5-15%) separating gel polymerization. Electrophoresis being carried out with an ascending voltage of 80V for the stacking gel and with 120V (2.5 hours) -150V (2.5 hours) -180V(1 hour) for the separating gel efficiently prevented band distortion and protein diffusion.

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