Question

You are interested in studying potential interaction of two proteins in cultured cells. What technique would best accomplish this goal? Include a description of how this technique works in your answer, and the possible results that would be expected (for a positive interaction or conversely, no interaction). (4 pts) You are starting a new research project and are interested in determining the function of a novel gene in cultured cells (called novl). What technique would you employ to study the function of novl in cultured cells? Include a description of the technique and any major steps involved in your answer (note that there is more than one possible technique that could be used here but only one should be included) (5 pts)

Answer 1

Answer:

1. One of the classic method to study protein-protein interactions is co-immunoprecipitation. In this method, you isolate or precipitate the protein of interest with a specific antibody. This is followed by studying the interactions with other protein of interest and analyzing their interactions through western blotting. You blot your antibody interactions with the second protein antibody. Hence a positive interaction shows the band while no interactions leads to no band in western blotting technique.

2. To determine the function of novel gene:

Realtime PCR : The technique helps in identifying the altered expression levels of the nov1 gene in cultured cells. The altered expression can be either increase or decrease in cells. This is one of the transcriptomics study performed to study the novel gene functions. others are northern blotting and DNA microarrays. For analysis, you culture the cells, isolate RNA, reverse transcribe to cDNA, and then perform real time PCR analysis with positive nad negative controls. Other approaches are gain and loss of function of the gene and cloning the gene.