Open AccessTechniques for rapid preparation of tomato leaf DNA and its application in real-time quantitative PCR-based transgene detection
Author(s) -
Weiwei Wang,
Changqing Zhu,
Xiaohua Liu,
Kunsong Chen,
Changjie Xu
Publication year - 2011
Publication title -
yichuan
Language(s) - English
Resource type - Journals
ISSN - 0253-9772
DOI - 10.3724/sp.j.1005.2011.01017
Subject(s) - solanum , dna extraction , dna , real time polymerase chain reaction , chromatography , centrifugation , biology , transgene , extraction (chemistry) , polymerase chain reaction , volume (thermodynamics) , microbiology and biotechnology , chemistry , gene , horticulture , biochemistry , physics , quantum mechanics
Using tomato (Solanum lycopersicum L. cv. Micro-Tom) leaf as material, a simple and rapid DNA preparation protocol was established. This method required only 2-20 mm2 leaf with only one extraction solution and involved one pipetation and one centrifugation each. No precipitation was required. The suitable volume of prepared DNA solution, as PCR template, for real-time quantitative PCR was determined to be 0.10.2 μL in 12.5 μL final reaction volume. The excessive template DNA solution was confirmed to reduce PCR efficiency and even can result in PCR failure. This technique for rapid preparation of DNA and a compatible real-time quantitative PCR were successfully applied in transgene detection of tomato plants.