Cloning and High Expression Anabaena Class-II Fructose-1, 6-bisphosphate Aldolase Gene in Escherichia coli / 中国生物工程杂志

The complete genomes of more cyanobacterial strains have been completed for sequence, and genetic engineering of cyanobacteria has evolved in the post-genome era. Since Kaneko and colleagues had completed the sequence for the complete genome of Anabaena sp. PCC 7120 in 2001, functions of some genes in this genome, including fructose-1,6-bisphosphate aldolase (FBA) gene, have been predicated using the method of bioinformatics. However, little information is available regarding whether this gene can encode FBA and its product characteristics of related enzyme. Here, to explore this information, the predicted II-FBA gene-encoding region in Cyanobase database was cloned by PCR method and then ligated into pET-32a to generate the expression vector, pET-FBA-II. The results of SDS-PAGE indicated that the expression level of the expected target polypeptide was approximate 23.4 percent as compared to total protein and the molecular weight is about 40 kDa as compared to the protein molecular marker. The results of enzyme activity analysis showed that the activity of II-FBA was ~11.8 U per mg protein and owned a standard activity of II-FBA. To sum up, the results not only prove the functional prediction of this II-FBA gene from the Cyanobase database, but also provide the important conditions for further studying its physiological and biochemical characteristics and functions of the gene expression product.

Cloning and Transformation of lba Gene of Glycine max into Chloroplast of Chlamydomonas reinhardtii / 中国生物工程杂志

To decrease the oxygen content in the cell is a key method to improve hydrogen production in Chlamydomonas reinhardtii.A new approach was developed by transforming the leghemoglobin gene lba,which has high affinity to oxygen,into the chloroplast of C.reinhardtii to get a low dissolved oxygen in the cell and result into improvement of H2 ase activity and H2 yield. The results showed that lba was successfully transformed into the chloroplast of C.reinahrdtii strain 849 and did not affect its growth significantly. The work paved the road for further regulation of lba expression in the chloroplast to improve of hydrogen production of C.reinahrdtii.