Question

1- How do you set up a restriction digest? What are the controls?   What is the purpose of controls? What are the components and what is the purpose of each component?

2- Why is mixing a critical step?

3- What kinds of information can you get by examining your phage by electron microscopy?

4- What is centrifugation? How does it work? Why is it useful? What properties of molecules can be exploited to achieve separation of a mixture of molecules in a suspension by centrifugation?

Answer 1

1 To setup a Restriction setup a Restriction digestion Experiment the up as protocol is followed 1. A Reaction minture is prepared by Components adding in the the following desired amount. Experimental Control ONA (to be digested) buffer (provide ambient pH to RE) DNA ( to be digested) buffer Restriction Enzyme (RE) Nucleuse free H₂O • Nuclease fure H₂O. Respiction Enzyme is not added." 2. Incubate the reaction at digestion temperature a 39°c. for 2 howy. 3. stop the digestion by heating or by EDTA Either. Controls help to or determine occured wether has igestion the digestion has a not. 2. Mixing helps the Restriction Enzymes to work efficiently. 3. Analyzing a phage by electron microscopy helps to examine the virus at molecular reuel . Eg capsid of tailed me phages is monophyletic in Origin - Capsid arrangement. 4- Centrifugation is a process used to separate or concentrate materials suspended in a liquid medium. It is a method to separate molecules based on their sedimentation rate under centrifugal field. It involves the use of the centrifugal force for the sedimentation of molecules. & It is used to measure such as molecular physical properties weight, density and shape. '