You have attempted to ligate a 1.5 kb fragment of foreign DNA into the EcoRI site in the multiple cloning site of the 4.0 kb plasmid vector shown below:
‪ (a) After ligation you use the DNA in the ligation mixture to transform host bacteria. Why is it important to use host bacteria that are deficient for restriction-modification?
(b) You screen the bacteria that supposedly have been transformed with recombinant plasmid DNA. Some of the bacterial colonies growing on the nutrient agar plate that contains ampicillin and X-gal are white and some are blue. Explain these results.
(c) To confirm the presence of the foreign DNA insert, you perform EcoRI restriction endonuclease digests on DNA extracted from bacterial colonies. On the diagram of an agarose gel shown below, draw the pattern of bands (label their size in kb) you would expect to see for EcoRI-digested recombinant plasmid and EcoRI-digested non-recombinant plasmid vector, after electrophoresis and staining of the gel with ethidium bromide.

Question

You have attempted to ligate a 1.5 kb fragment of foreign DNA into the EcoRI site in the multiple cloning site of the 4.0 kb plasmid vector shown below:
‪  (a) After ligation you use the DNA in the ligation mixture to transform host bacteria. Why is it important to use host bacteria that are deficient for restriction-modification?
(b) You screen the bacteria that supposedly have been transformed with recombinant plasmid DNA. Some of the bacterial colonies growing on the nutrient agar plate that contains ampicillin and X-gal are white and some are blue. Explain these results.
(c) To confirm the presence of the foreign DNA insert, you perform EcoRI restriction endonuclease digests on DNA extracted from bacterial colonies. On the diagram of an agarose gel shown below, draw the pattern of bands (label their size in kb) you would expect to see for EcoRI-digested recombinant plasmid and EcoRI-digested non-recombinant plasmid vector, after electrophoresis and staining of the gel with ethidium bromide.

Quizplus · Accepted Answer

The Answer of You have attempted to ligate a 1.5 kb fragment of foreign DNA into the EcoRI site in the multiple cloning site of the 4.0 kb plasmid vector shown below:
‪  (a) After ligation you use the DNA in the ligation mixture to transform host bacteria. Why is it important to use host bacteria that are deficient for restriction-modification?
(b) You screen the bacteria that supposedly have been transformed with recombinant plasmid DNA. Some of the bacterial colonies growing on the nutrient agar plate that contains ampicillin and X-gal are white and some are blue. Explain these results.
(c) To confirm the presence of the foreign DNA insert, you perform EcoRI restriction endonuclease digests on DNA extracted from bacterial colonies. On the diagram of an agarose gel shown below, draw the pattern of bands (label their size in kb) you would expect to see for EcoRI-digested recombinant plasmid and EcoRI-digested non-recombinant plasmid vector, after electrophoresis and staining of the gel with ethidium bromide.

You Have Attempted to Ligate a 1