[Optimization of reaction conditions for RAPD analysis of freshwater planarians in China].
Using the Genomic DNA purification kit, the total DNA of the freshwater planarian was extracted and developed one single band through 0.8% agarose gel electrophoresis with OD260/OD280 between 1.5 and 2.2, which could satisfy the requirements of RAPD and PCR on DNA. With the extracted DNA template, we tested experimental conditions that might affect RAPD results including annealing temperature, concentrations of template DNA, primer, Mg2+ and dNTPs. Through comparision we found that it was necessary and important to optimize the experimental conditions for producing stable and repeatable RAPD results. The optimized reaction conditions of RAPD for freshwater planarian in 25 microl reaction volume were as follows: 20 ng template DNA, 37 degrees C annealing temperature, 0.2 micromol/L primer, 2.0 mmol/L Mg2+ and 200 micromol/L dNTPs.
Zhang
He Cai
HC
College of Life Sciences, Henan Normal University, Xinxiang 453002, China.
Chen
Guang Wen
GW
Li
Yu Chang
YC
Xu
Cun Shuan
CS
D000818:Animals; D002681:China; D017931:DNA Primers / Q000235:genetics; D018509:DNA, Helminth / Q000235:genetics; D004587:Electrophoresis, Agar Gel; D005618:Fresh Water; D010932:Planarians / Q000235:genetics*; D016133:Polymerase Chain Reaction; D019105:Random Amplified Polymorphic DNA Technique; D013696:Temperature
D017931:DNA Primers; D018509:DNA, Helminth