The DNA Extraction from Chlorella Algae, and its Preparation for Real Time-polymerase Chain Reaction

Abstract

Our current research involved the extraction of DNA from chlorella algae for prepare it to real-time polymerase chain reaction (RT-PCR) After the collection of algal samples from the riverine quarter environment, they were placed in test tubes, about (1-mL) and centrifuged at (3500 rotation for 5 minutes), with placed a control negative sample for certain the success of the practical steps sequentially of the experiment, and to know the effectiveness of the lytic enzymes, and the reagents which used. When the microalgae are visible down each test tube by watching its green color, 400 μL of the lytic solution add to the remaining sample with mixing, and 10 μL of an enzyme proteinases K are added to them, which helps to break down the walls and membranes of the cells. The incubated of samples at a temperature of 65°C for 1-hour, followed by the addition 400 μL of chloroform phenol (it plays the role of breaking down DNA-related proteins) with shaking, and the samples are centrifuged again to take the upper layer as it represents the DNA of the algal samples, they are withdrawn and transferred to a new pentroph. A 5 μL RNA-ase enzyme is added to them to destroy the RNA and preserve the DNA. Added (500 mL of isopropanol), with mixing well by hand, and centrifuged at 12,000 rotation, for 10 minutes, alcohol removed and DNA extracted prepared for the PCR reaction.