Question

Which of the following is important for the design and use of a reporter GFP fusion protein plasmid to study the subcellular localization of a protein of interest? Select all the apply. Question 9 options:

the STOP codon from the gene of the protein of interest must be removed before insertion into the MCS upstream of the GFP gene.

The GFP gene should be downstream of the promoter of the gene of interest.

The GFP gene should be in the same reading frame as the gene of interest.

The construct with the GFP gene and the gene of the protein of interest must be transduced into cells.

Answer 1

The stop codon from the gene of the protein of interest must be removed before insertion into the MCS upstream of the GFP protein because without this the stop codon would lead to an end of transcription and translation after the desired genome and GFP wouldn't be translated hence won't be expressed.

The GFP gene should not be downstream of the promoter of the gene of interest because that may lead to expression of ONLY GFP but not the desired gene ,which will ultimately lead to error.

The GFP gene should be in the same reading frame as the gene of interest so that they will be transcripted and translated together.

The construct with the GFP gene and the gene of the protein of interest are generally ligated into a vector and then transformed or transduced into the cells.

So here option 1 and 3 are the right answers. Give a thumbs up if you like the answer. Thank you