An improved chloramphenicol acetyltransferase assay for Plasmodium falciparum transfection.
Chloramphenicol acetyltransferase (CAT) is a popular choice as a reporter gene in transgenic studies in many different organisms, including Plasmodium falciparum. For experimental investigations into transfection efficiency and gene expression a robustly quantitative assay is of great value. On investigation the published protocol for CAT assay of P. falciparum was found to be prone to saturation due to the long incubation time; moreover, cellular material extracted from the parasite increased the enzyme activity. A new protocol was developed which is quantitative across a range of three orders of magnitude of CAT activity, takes account of the cellular extract effect, and is more rapid than the established method. The value of these improvements was demonstrated by analysing the effects of parasitaemia and amount of plasmid on transfection efficiency with both old and new methods.
Lucas
Stuart J
SJ
Division of Parasitology, National Institute for Medical Research, The Ridgeway, Mill Hill, London NW71AA, UK. stu_nami@yahoo.co.uk
Holder
Anthony A
AA
D000818:Animals; D015500:Chloramphenicol O-Acetyltransferase / Q000032:analysis* / Q000235:genetics*; D004912:Erythrocytes / Q000469:parasitology; D015870:Gene Expression; D017930:Genes, Reporter; D006801:Humans; D066298:In Vitro Techniques; D016778:Malaria, Falciparum / Q000469:parasitology; D010963:Plasmodium falciparum / Q000201:enzymology* / Q000235:genetics* / Q000254:growth & development; D011994:Recombinant Proteins / Q000032:analysis / Q000235:genetics; D014162:Transfection
D011994:Recombinant Proteins; D015500:Chloramphenicol O-Acetyltransferase