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A PCR amplification method without DNA extraction
Author(s) -
Li Hongwei,
Xu Haiyue,
Zhao Chunjiang,
Sulaiman Yiming,
Wu Changxin
Publication year - 2011
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.201000392
Subject(s) - tris , hydroxymethyl , ethylene diamine , chromatography , reagent , lysis , chemistry , buffer (optical fiber) , lysis buffer , proteinase k , dna , dna extraction , extraction (chemistry) , buffer solution , polymerase chain reaction , microbiology and biotechnology , biochemistry , biology , nuclear chemistry , telecommunications , computer science , gene , stereochemistry
To develop a simple and inexpensive method for direct PCR amplification of animal DNA from tissues, we optimized different components and their concentration in lysis buffer systems. Finally, we acquired the optimized buffer system composed of 10 mmol tris(hydroxymethyl)aminomethane (Tris)‐Cl (pH 8.0), 2 mmol ethylene diamine tetraacetic (EDTA) (pH 8.0), 0.2 mol NaCl and 200 μg/mL Proteinase K. Interestingly, the optimized buffer is also very effective when working with common human sample types, including blood, buccal cells and hair. The direct PCR method requires fewer reagents (Tris‐Cl, EDTA, Protease K and NaCl ) and less incubation time (only 35 min). The cost of treating every sample is less than $0.02, and all steps can be completed on a thermal cycler in a 96‐well format. So, the proposed method will significantly improve high‐throughput PCR‐based molecular assays in animal systems and in common human sample types.