Devise an experiment, based on sucrose gradient sedimentation, that would demonstrate that proteins are synthesized on polyribosomes.

To devise an experiment using sucrose gradient sedimentation to demonstrate that proteins are synthesized on polyribosomes, follow these steps:

1. Cell Preparation: Start by culturing cells (such as bacterial or mammalian cells) and harvesting them at a specific time point during active protein synthesis.

2. Cell Lysis: Lyse the cells carefully using a suitable lysis buffer to release all cellular components, including polyribosomes bound to mRNA.

3. Centrifugation: Centrifuge the cell lysate at a low speed (around 10,000 × g) to remove cell debris and obtain a homogenized supernatant containing ribosomes.

4. Sucrose Gradient Preparation: Prepare a sucrose gradient by layering different concentrations of sucrose in a centrifuge tube. Higher sucrose concentrations are at the bottom, while lower concentrations are at the top. Typically, a gradient ranging from 5% to 50% sucrose can be used.

5. Loading the Gradient: Carefully load the cleared supernatant from step 3 onto the top of the sucrose gradient.

6. Ultracentrifugation: Centrifuge the sucrose gradient at high speed (e.g., 100,000 × g) for several hours. This process separates the polyribosomes based on their sedimentation rate in the sucrose gradient.

7. Fraction Collection: Collect fractions from the bottom of the centrifuge tube using a fraction collector or by carefully extracting fractions with a syringe. It is essential to collect fractions slowly from the bottom without disturbing the gradient.

8. Protein Analysis: Analyze the collected fractions for the presence of proteins using methods like SDS-PAGE or Western blotting. Select fractions that appear enriched in protein content for further analysis.

9. RNA Analysis: Extract RNA from the chosen protein-enriched fractions and perform reverse transcription-polymerase chain reaction (RT-PCR) or other mRNA analysis techniques to confirm the presence of mRNA co-sedimenting with the polyribosomes.

10. Control Experiment: Perform a control experiment using a treatment that disrupts polyribosome formation, such as the addition of puromycin, which causes premature translation termination. This control helps to demonstrate that only polyribosomes, not individual ribosomes, are responsible for protein synthesis.

By comparing the distribution of proteins and mRNA in the sucrose gradient fractions with and without the control treatment, you can confirm that proteins are synthesized on polyribosomes.