An immunochemical method for mRNA purification. Application to messenger RNA encoding trypanosome variable surface antigen.
A simplified procedure for isolation of specific mRNA using Staphylococcus aureus protein A immunoadsorbent chromatography has been developed. This procedure has been applied to the mRNA encoding Trypanosoma brucei variable surface antigen. Trypanosome polyribosomes were reacted with antibodies isolated from an anti-variable antigen-specific serum by protein A-Sepharose column chromatography. Antibody-bound variable antigen-synthesizing polyribosomes were then separated from unbound polyribosomes also by protein A-Sepharose column chromatography. This simple protocol gave a greater than 50% yield of variable antigen-specific mRNA which appeared to be very highly purified as determined by translation in an mRNA-dependent reticulocyte lysate assay. The mRNAs encoding three different T. brucei surface antigens have been purified. The procedure described here should be useful in purification of other mRNAs.
D000818:Animals; D000954:Antigens, Surface / Q000235:genetics*; D008970:Molecular Weight; D011132:Polyribosomes / Q000378:metabolism; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics / Q000302:isolation & purification*; D011817:Rabbits; D012156:Reticulocytes / Q000378:metabolism; D014346:Trypanosoma brucei brucei / Q000378:metabolism*
D000954:Antigens, Surface; D012333:RNA, Messenger