QUESTION 8 A scientist carries out a ligation reaction designed to insert a fore

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QUESTION 8 A scientist carries out a ligation reaction designed to insert a foreign piece of DNA into a plasmid that contains the front end of the lacZ gene within the multiple cloning site (MCS). He carries out three transformations in parallel with lacZ bacteria. The three transformations (i, ii, and ii) contain the following i. sterile water ii. a sample of the original plasmid ii. the ligation reaction He then plates the transformations on medium containing X-gal and ampicillin Reference: Ref 19-3 What results would indicate the scientist should proceed to culture colonies from (iii) that might contain the desired clone? O A. No colonies on plate (i), blue colonies on plate (ii), and predominantly blue colonies on plate (ili). B. No colonies on plate (i), white colonies on plate (i), and predominantly blue colonies on plate (ii) C. No colonies on plate (i), blue colonies on plate (i), and predominantly white colonies on plate (ii) D. Many blue colonies on all three plates E. White colonies on plates (i) and (i) and predominantly blue colonies on plate (ii) QUESTION 9 A scientist edits a mammalian genome by inserting a GFP reporter sequence downstream of a gene of interest. Which of the following methods is she likely to have used to make this change? O A. oligonucleotide-directed mutagenesis B. site-directed mutagenesis O C. CRISPR-Cas9 genome editing followed by nonhomologous end joining (NHEJ) D.mutagenesis with radiation O E. CRISPR-Cas9 genome editing followed by homologous recombination (HR) with a donor template

Explanation / Answer

8. Option C is correct.

Insertional inactivation: Insertion of the foreign DNA into the MCS of the vector disrupts the coding sequence of a reporter gene such as GUS or GFP. So, true transformants can not produce the blue color in the presence of X-gal.

Sterile water should not produce any colonies as it does not contain DNA.

A sample of original plasmid should produce only blue colonies as the GUS reporter CDS is intact in this plasmid.

The ligation sample should produce predominantly white colonies as the GUS reporter CDS is disrupted in this plasmid.

9. Option E is correct.

To create insertional mutants with a reporter gene, CRISPR-Cas9 genome editing method followed by HR with the donor template is used.

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