Difference between revisions of "Part:BBa K2904011"

Revision as of 08:54, 20 October 2019

Modular Adda riboswitch containing Tuner B

Design

Background of 2019 OUC-China's project——RiboLego

Due to context-dependent performance and limited dynamic range, the widespread application of riboswitches is currently restricted. By replacing its original ORF with a new one, the structure of an aptamer domain can be subtly disrupted, resulting in a loss of ligand response. So riboswitch is still not be considered as a ‘plug and play' device. To tackle these problems, our project focuses on a standardized design principle to be used for modular and tunable riboswitch. The modular riboswitch we defined consists of the original riboswitch, Stabilizer and Tuner. Stabilizer can protect the structure of riboswitch from damage while Tuner can reduce the expression probability of fusion protein and make improvement of riboswitch function.

The construction of this part

To validate our design guideline, we employed a activating Adda riboswitch, which can regulate the expression of adenosine deaminase by binding 2-aminopurine in Vibrio vulnificus.The first 150bp of adenosine deaminase was chosen as Stabilizer of Adda riboswitch because our docking matrix suggested that a normal riboswitch structure would be observed when using this length of Stabilizer. This part was modular Adda riboswitch containing the original riboswitch, Stabilizer and Tuner B.

Result

In order to confirm the function of modular Adda riboswitch containing Tuner B, we tested our system by microplate reader, which is used to reflect the intensity of sfGFP changing over time. The following chart shows the dynamic curve measured every two hours. It can prove that the modular Adda riboswitch can restore the structure of riboswitch and control the downstream gene expression during the whole cultivation period.

Figure1: The results of modular Adda riboswitch containing Tuner B by microplate reader.

Sequence and Features