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ID: 148874 • Letter: U
Question
USING THIS ARTICLE LINK: http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0001-37652015000200691&lng=en&tlng=en
1- What is the major idea of the paper? Which technique gave the best result? (2 points)
2- How did they get rid of the soil in the samples (before testing the 4 techniques)? Were they able to get rid of all the soil in the samples (again before testing the 4 techniques)? (3 points)
3- When the DNA was extracted some methods did not lead PCR products. Why was that? (3 points)
Explanation / Answer
1.The major idea of the paper was to find out which is an efficient way to extract DNA of the different nematode communities that are present in the soils of Argentine Pampas agricultural area out of the four DNA isolation techniques and then carry out PCR amplification of the DNA.The CTAB method that was combined with proteinase K and phenol-chloroform-isoamyl alcohol gave the best result for DNA extraction.
2.They got rid of the soil in the samples by homogenizing the sample in a magnetic shaker and collecting the sub samples of 5 ml from it from which DNA was extracted. No, they were not able to get rid of all the soil in the samples before testing the 4 techniques as humus and humic acid particles from soil were still present.
It was during the application of the 4 techniques that soil that was still present was removed.In two different extraction methods, Chelex resin and CTAB with phenol chloroform were used.Chelex resin disperses the soil particles and CTAB functions to remove humus materials ,humus acids that present in the soils and which inhibit the functioning of PCR enzymes.Phenol chloroform helps in deproteinization of the homogenate formed and also helps to remove humus and pigments present in soil.Through CTAB-phenol chloroform technique,all soil could be got rid of and results were clear and good but not when Chelex resin was used.
3.Some DNA extracts did not lead to PCR products as soil particles containing humus and humic acid were still present in the sample and these acted as inhibitors for PCR enzymes; specially the Taq polymerase enzyme.
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