Question

1.How are the contents of the PCR tube analyzed to determine a positive or negative result?

2. What does a "positive" PCR result look like?

What dies a "Negative" PCR result look like?

Answer 1

1.

The reaction mixture of PCR contains -

Template DNA, pair of primers, dNTPs, taq polymerase, magnesium chloride, buffer and water.

Negative control does not contain template DNA.

Positive control contains template DNA which will for sure be amplified by the given pair of primers.

The importance of negative control is to ensure that there is no mispriming and the primer binds to only the template DNA. The taq polymerase does not use primer and deoxyribonucleotides to synthesise a new copy of daughter DNA molecule by itself. In this case there will be no amplification and no formation of PCR product.

The importance of positive control is to ensure that the primers which we have used are functional and they will bind to a given template DNA if there is a complementary region between primer and temperate DNA. In this case there will be amplification for sure and we will get a PCR product.

The experimental tube is the one which contains all the reagents and the template DNA contains the target region. In this case the amplification and formation of PCR product depends upon the interaction between all the region including primer and template DNA.

2. If we run the product obtained after PCR on gel electrophoresis, then we will see a band in case of positive PCR and there will be no band in case of negative PCR because amplification has not occurred.

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