Purification of immunoglobulin heavy chain messenger RNA by immunoprecipitation from the mouse myeloma tumor, MOPC-31C.
Immunoglobulin heavy chain mRNA was purified from immunoprecipitated polysomes derived from the mouse myeloma tumor, MOPC-31C. The purified mRNA migrated predominantly as a single band upon polyacrylamide gel electrophoresis in 98% formamide and the molecular weight of this mRNA was calculated to be 700,000. This mRNA was as active as the purified light chain mRNA when it was employed as a template in a cell-free protein synthesizing system from wheat germ. The translation product had a molecular weight of 55,000 daltons, and migrated slightly faster than mature heavy chain upon polyacrylamide gel electrophoresis in sodium dodecylsulfate. The protein synthesized by the direction of this mRNA was shown to yield tryptic peptides corresponding to those derived from the mature heavy chain protein except that one missing peptide was replaced by another additional peptide. DNA complementary to the mRNA was synthesized by RNA-dependent DNA polymerase from avian myeloblastosis virus. Hybridization kinetic analysis between the heavy chain mRNA and its complementary DNA indicated that the RNA was essentially homogenous with rabbit globin mRNA as a standard.
D000818:Animals; D009189:Avian Myeloblastosis Virus / Q000201:enzymology; D002478:Cells, Cultured; D004247:DNA / Q000096:biosynthesis; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D007143:Immunoglobulin Heavy Chains / Q000096:biosynthesis*; D007930:Leucine / Q000378:metabolism; D051379:Mice; D008970:Molecular Weight; D009101:Multiple Myeloma / Q000032:analysis*; D009194:Myeloma Proteins / Q000096:biosynthesis; D009691:Nucleic Acid Denaturation; D009693:Nucleic Acid Hybridization; D010446:Peptide Fragments / Q000032:analysis; D011132:Polyribosomes; D012333:RNA, Messenger / Q000302:isolation & purification* / Q000378:metabolism
D007143:Immunoglobulin Heavy Chains; D009194:Myeloma Proteins; D010446:Peptide Fragments; D012333:RNA, Messenger; D004247:DNA; D004259:DNA-Directed DNA Polymerase; D007930:Leucine