pcDNA3-Myc-His-Rap80 was used as the template to perform the in vitro translation (IVT) experiment with 35S-Met using the TnT Quick Coupled Transcription/Translation System (Promega) according to the manufacturer’s instructions. Two hundred microliters of 0.5 μM biotin-ds70mer DNA was incubated with 1 μg of K63-linked polyubiquitin (2 to 7) chains at room temperature for 2 hours and washed thoroughly to eliminate nonspecific binding. Increased amounts of IVT-Rap80 (10, 20, and 30 μl, respectively) were added and further incubated at 4°C for 2 hours before being washed thoroughly and resolved on SDS-PAGE for autoradiography.

Note: The content above has been extracted from a research article, so it may not display correctly.

Please log in to submit your questions online.
Your question will be posted on the Bio-101 website. We will send your questions to the authors of this protocol and Bio-protocol community members who are experienced with this method. you will be informed using the email address associated with your Bio-protocol account.

We use cookies on this site to enhance your user experience. By using our website, you are agreeing to allow the storage of cookies on your computer.