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SITE DIRECTED MUTAGENESIS ,[object Object]
[object Object]
[object Object],[object Object]
Basic Mechanism of Site-directed mutagenesis + polymerase + primer replication Mutant translation Wild type translation Only one amino acid changed Wild type protein Mutant protein primer (1) (2) (3) (5) (4) (6) Val -> Thr Smith (1993) Juang RH (2004) BCbasics CAG GTC CAG G C C CAG G C C CAG G C C C G G Thr GTC CAG Val
 
Primer design for site-directed mutagenesis ,[object Object],[object Object],[object Object],[object Object],[object Object],[object Object],5’ 5’ 3’ 3’ * *
TYPES OF SDM ,[object Object],[object Object],[object Object],[object Object]
Oligo-NT directed mutagenesis
SDM using PCR
Cassette Mutagenesis ,[object Object],[object Object],[object Object]
Random Chemical Mutagenesis ,[object Object],[object Object],[object Object],[object Object]
Chemical Target Na bisulphite Nitrous acid Formic acid Hydrazine dC dT dC Du dA dHX Glycosyl bonds break. Pyrimidine ring break.
Applications of SDM ,[object Object],[object Object],[object Object],[object Object],[object Object]
Papers and useful websites ,[object Object],[object Object],[object Object],[object Object],[object Object]
Thank You haris.p mac ernakulam

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SITE DIRECTED MUTAGENESIS.HARIS

  • 1.  
  • 2.
  • 3.
  • 4.
  • 5. Basic Mechanism of Site-directed mutagenesis + polymerase + primer replication Mutant translation Wild type translation Only one amino acid changed Wild type protein Mutant protein primer (1) (2) (3) (5) (4) (6) Val -> Thr Smith (1993) Juang RH (2004) BCbasics CAG GTC CAG G C C CAG G C C CAG G C C C G G Thr GTC CAG Val
  • 6.  
  • 7.
  • 8.
  • 11.
  • 12.
  • 13. Chemical Target Na bisulphite Nitrous acid Formic acid Hydrazine dC dT dC Du dA dHX Glycosyl bonds break. Pyrimidine ring break.
  • 14.
  • 15.
  • 16. Thank You haris.p mac ernakulam

Notas do Editor

  1. 定點突變 是使用人工合成的引子，先與目標基因雜合 (2) ，而此引子的核苷酸序列與目標基因互補，但其上有一個核苷酸變異 (GTC->G C C) 。然後補滿雙股核酸 (3) ，再把此混成基因轉殖進入宿主，則宿主菌的複製系統將會因複製而產生兩種質體 (4) ，其一為原來的野生型，轉譯可得原來的蛋白質 (5) ；另一則為突變型，轉譯則得到突變的蛋白質，但只改變了一個指定位置的胺基酸 (6) 。