A novel approach for extraction of PCR-compatible DNA from activated sludge samples collected from different biological effluent treatment plants.
This paper describes a method that facilitates the extraction of PCR-compatible DNA from different activated sludge samples. The approach involves a novel preprocessing step in DNA extraction, which removes potential PCR inhibitors. The sludge was washed with different ratios of acetone and petroleum ether after pretreatment with 0.01% Tween-20 at 50 degrees C. It was observed that an initial washing step with 50 mM Tris-HCl, pH 9.0, before the detergent-solvent step, improved the quality of the extracted DNA. The extraction protocol resulted in amplifiable amounts of DNA when 10 mg of a sludge sample was used, even in the presence of phenol as a sludge contaminant. The usefulness of the extracted template was demonstrated by carrying out different PCR reactions. The random amplified polymorphic DNA (RAPD) patterns demonstrated the diversity of sludge samples.
Purohit
Hemant J
HJ
National Environmental Engineering Research Institute, Nehru Marg, Nagpur 44 0020, India. hemantdrd@hotmail.com
Kapley
Atya
A
Moharikar
Aditi A
AA
Narde
Gurpreet
G
D004269:DNA, Bacterial / Q000032:analysis* / Q000302:isolation & purification*; D006863:Hydrogen-Ion Concentration; D007220:Industrial Waste; D016133:Polymerase Chain Reaction / Q000379:methods; D019105:Random Amplified Polymorphic DNA Technique / Q000379:methods*; D012722:Sewage / Q000382:microbiology*
D004269:DNA, Bacterial; D007220:Industrial Waste; D012722:Sewage