<i>Escherichia coli </i>cell line with <i>thyA </i>knockout and <i>folA </i>knockout

摘要

The objective of this invention is to create a double thyA folA knockout Escherichia coli (E. coli) strain for antifolate screening against DHFR of malaria and other parasites. This strain is used together with a plasmid expressing DHFR-TS from the desired pathogenic organism, which constitutes an anti-DHFR assay against the pathogenic organism of interest. The benefit of this invention is that there is no interference from either host DHFR or trimethoprim, a bacterial DHFR inhibitor. This tool is easy to use and maintain. It provides quick and reliable results as compared with conventional anti-malarial and anti-parasitic assays. This invention should facilitate discovery of new anti-DHFR compounds against malaria and other parasitic diseases.

主权项

1. An Escherichia coli (E. coli) cell line in which its thymidylate synthase (thyA) and dihydrofolate reductase (folA) genes have been disrupted by genetic knock out of the thy A and folA genes (E. coli thyA folA KO), wherein antibiotic resistance genes used to create the genetic knockout have been functionally eliminated, and wherein said cell comprises a parasite dihydrofolate reductase-thymidylate synthase (DHFR-TS) gene for heterologous expression of a parasite DHFR-TS enzyme.