Question

1) What is the purpose of incubating the bacteria with the digestive buffer?

2. Is the 100 degree C water bath used to denature proteins or DNA?

3. At which temperature do the primers bind to the target DNA during the PCR?

4. For the PCR what is absent from the negative control tube that should be in positive control tube and sample tube?

5. What is in the mixture of second PCR prior to DNA sequencing that is not in the first PCR mixture?

Answer 1

Question 1:

What is the purpose of incubating the bacteria with the digestive buffer?

Answer:

The purpose of incubating the bacteria with the digestive buffer is the lysis of the bacteria. Due to incubation in digestive buffer lysis of bacterial cell wall will occur and the DNA content comes out which can used in experiment of molecular biology.

Question 2:

Is the 100 degree C water bath used to denature proteins or DNA?

Answer:

Yes, the 100 degree C water bath is used to denature proteins or DNA. If a DNA is heated in 100 degree C hot water bath for longer time it will undergo denaturation. On the other hand if proteins are heated in 100 degree C hot water bath for longer time it will also undergo denaturation.

Question 3:

At which temperature do the primers bind to the target DNA during the PCR?

Answer:

Binding of the primers to the target DNA during the PCR is known as annealing step. In this step the primers bind to the target DNA at about 50-55 degree centigrade temperature.

Question 4:

For the PCR what is absent from the negative control tube that should be in positive control tube and sample tube?

Answer:

For the PCR in the negative control tube template DNA is absent that should be in positive control tube and sample tube. In negative control tube all the requirements i.e., dNTP mix, forward primer, reverse primer, Taq polymerase and suitable buffer are added but no template strand is added. The negative control in PCR is used to check whether there is any contamination or not in the used buffer solutions. If any amplification takes place, this will mean that the solution used is contaminated. This will help to solve any issues related to false positive amplification. On the other hand in positive control tube and sample tube template DNA is present. In positive control tube known template DNA is used to control the experiment and in sample tube unknown template DNA is used which is determined by the PCR experiment.