Efficient in vitro expression of interferon alpha analogs using SP6 polymerase and rabbit reticulocyte lysate.
We have investigated the use of in vitro expression as a quick and convenient means of screening large numbers of interferon (IFN) analogs generated using in vitro mutagenesis. The IFN-alpha 1 mRNA generated from DNA template using SP6 RNA polymerase is efficiently translated in rabbit reticulocyte lysate (RRL). The antiviral specific activity of this RRL-synthesized IFN-alpha l is equivalent to the yeast-synthesized protein. In contrast with the yeast-expression system, where some IFN-alpha analogs are poorly expressed, all analogs tested were well expressed in RRL.
Tymms
M J
MJ
Centre for Molecular Biology and Medicine, Monash University, Clayton, Victoria, Australia.
McInnes
B
B
D000818:Animals; D001681:Biological Assay; D002474:Cell-Free System; D003001:Cloning, Molecular; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005786:Gene Expression Regulation; D007370:Interferon Type I / Q000235:genetics*; D008715:Methionine / Q000378:metabolism; D009154:Mutation; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000096:biosynthesis; D011817:Rabbits; D012156:Reticulocytes / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics; D014158:Transcription, Genetic
D007370:Interferon Type I; D012333:RNA, Messenger; D008715:Methionine; D012321:DNA-Directed RNA Polymerases