Team:XJTLU-CHINA/Parts Overview

Part Overview

This year, the parts we designed can be divided into 2 types: therapeutic parts and regulatory parts.

Therapeutic parts

We designed parts containing Excitatory Amino Acid 2 (EAAT2) coding sequence. The expression of EAAT2 proteins in neuroblastoma cells/astrocytes can efficiently absorb extracellular L-glutamate. Our designed part encoding EAAT2 sequence showed high compatibility to the mammalian cell lines we used (N2a cell lines).

Besides that, to achieve a high RNA packaging efficiency, we designed a C/D box region at the downstream of EAAT2 coding sequence. By this approach, untranslated EAAT2 coding mRNAs can specifically bind with CD63-L7Ae fusion protein, which is a previous design from XJTLU-CHINA 2018. This composite part can be placed on pcDNA3.1+ plasmids by normal standard assembly methods. The device consisting of EAAT2 coding sequence, C/D box and pcDNA3.1+ backbone has achieved a great results in inducing extracellular L-glutamate intake our experiments this year.

Regulatory parts

Our regulatory parts contains 2 categories: ligand-responsive ribozyme switch and regulation protein. For ligand-responsive ribozyme switches, they can be divided into 2 categories: protein-responsive ribozyme switches and theophylline-responsive ribozyme switches.

Protein-responsive ribozyme switches are designed to be applied in mammalian cell lines (eg. HEK293T), aims to build up genetic circuits in cells and further to control the expression of gene(s) temporally. To interact with these switches intracellularly, we designed regulation proteins corresponded to these protein-responsive switches. Each type of protein can specifically interact with riboswitches when they are expressed in host cells.

We also designed and submitted theophylline-responsive riboswitches which is compatible in prokaryotes such as E. Coli. These parts are designed to text the function of theophylline riboswitch sequences. Furthermore, we submitted our improved theophylline-responsive switch from PKU-R to achieve a better sensitivity to theophylline.