A direct method for cDNA cloning employing translation-released mRNA
Barardi, C.R.; Santos, C.L.; Villa, L.L.; Brentani, R.R.
Brazilian Journal of Medical and Biological Research 20(3-4): 313-320
1987
ISSN/ISBN: 0100-879X PMID: 3451785 Document Number: 295289
1. A new cloning procedure is described for cDNA synthesis from mRNA released by in vitro translation of polysomes in a cell-free amino acid incorporating system. The usefulness of the method lies in the feasibility of employing nanogram amounts of mRNA. 2. Complementary DNA is synthesized directly in the translation mixture simply by adjusting the concentration of some components and removing ribosomes by boiling and centrifugation. 3. As an example, we report here the construction and characterization of a cDNA clone corresponding to chick alpha(I)procollagen starting from a collagen-synthesizing polysome fraction obtained from chick embryos.