Question

You discovered a novel eukaryotic organism that glows in the dark. You believe this trait is due to a single gene, and you wish to clone the gene. which of the following strategies is most likely to be successful?

• A. Isolate the genomic DNA from the organism, digest with a restriction endonuclease, insert into a plasmid vector and transform into a plasmid
• B. Isolate the genomic DNA from the organism, digest with a restriction endonuclease, insert into a plasmid vector and transform into bacteria. Screen colonies for for the ability to glow in the dark.
• C. Isolate mRNA from the organism, reverse transcribe and generate cDNA, insert into a plasmid vector and transform into bacteria. Screen colonies for the ability to glow in the dark.
• D. Isolate mRNA from the organism, reverse transcribe and genetate cDNA, Insert into a plasmid vector and transform into eukaryotic cells such as yeast. Screen colonies for the ability to glow in the dark.

Answer 1

Answer: (D)

Isolate mRNA from the organism, reverse transcribe and genetate cDNA, Insert into a plasmid vector and transform into eukaryotic cells such as yeast. Screen colonies for the ability to glow in the dark.

As it is a eukaryotic gene, it is better to be incorporated in a eukaryotic cell. Digestion by restriction endonuclease is not possible as the gene is unidentified and is to be isolated and cloned. It is better and easier to identify the gene transcript, i.e., protein coding mRNA, reverse transcribe it to produce cDNA and then transferring this intron less cDNA into an eukaryotic cell like yeast through a plasmid vector.

So, the correct answer is 'Isolate mRNA from the organism, reverse transcribe and generate cDNA, Insert into a plasmid vector and transform into eukaryotic cells such as yeast. Screen colonies for the ability to glow in the dark'.