Question

3. You have now switched to yet another project focused on studying the mechanisms of splicing (you are one busy researcher!). You believe that you have identified a novel splicing factor (called NSF1) that is directly associated with the US snRNP and is critical to its function in the splicing reaction. What experiment could you perform to demonstrate that NSF1 and U5 snRNP interact with one another and are part of the same complex in cells? (5 pts) (Note: this question has more than one correct answer - whichever technique(s) you choose, be sure to completely describe the experiment and associated methods, as well as the expected results)

Answer 1

3.
Aim: To check whether NSF1 can associate with U5 snRNP
Experiment: Coimmunoprecipitation (Invitro pulldown)
Procedure: Incubate NSF1 with U5-snRNP in a suitable buffer. Use antibodies to pull down NSF1 and probe the sample with U5-snRNP. If both proteins interact, we should observe a band in the mixture.

Aim to check whether NSF-1 can associate with U5 snRNP experiment communopreciation invitro pulldown procedure incubate NSF1 with u5-snRNP in a suitable buffer use antibodies to pull down NSF1 and probe the sample with u5-snRNP if both proteins interact we should observe a band in mixture if both the proteins interact with each other banding pattern can be observed as below if both the proteins do not interact with each other banding pattern can be observed as below