Computer-aided design to raise the expression level of the M3-M4 loop in the essential subunit of human NMDA receptor in E. coli

To improve the expression of foreign gene in pBV220 vector, a model was established by predicting RNA secondary structure based on helical regions random stacking and calculating the codon adaptation. The model showed a statistically significant linkage between high (low) expression and the free energy of secondary structure in the regions -30--39 of 5^end and 30-- -39 of 3^end, while the local codon adaptation of 9 bp in the 3^end was also related to expression level. Guided by this model, a pair ofo PCR primers was designed with RNAfold and Goldkey software. The primers were used for expressing the M3-M4 loop gene in the essential subunit of human NMDA receptor which was amplified and ligated into pBV220 vector. By transforming DH5#alpha# with this recombinant plasmid, M3-M4 loop was highly expressed in DH5#alpha# transformant with expression level over 20%.