Isolationand in vitro translation of leghaemoglobin mRNA from yellow lupin root nodules.

Messenger RNA for leghaemoglobin, extracted from polysomes of yellow lupin root nodules, was purified by chromatography on oligo(dT1-cellulose followed by sucrose-gradient centrifugation. Leghaemoglobin mRNA activity was assayed in a cell-free protein synthesizing system derived from wheat germ. The purified mRNA was sedimented in surcrose gradient in the 9S region which corresponds to a molecular weight of about 225,000. The poly(A) segment was estimated to be 66 nucleotides in length, based on hybridization with [3H]poly(U). Analysis of the translation product using immunoprecipitation and polyacrylamide-gel electrophoresis in denaturing conditions revealed that the product made in vitro was identical with authentic leghaemoglobin.