RNA transcribed in vitro from myeloblast chromatin by exogenously added RNA polymerase B predominantly consists of short chains that remain in hybrid structure with the template; the remainder of the product is free RNA of heterogeneous size. Addition of polyanions during synthesis caused an increase in the size and amount of free RNA with a concomitant decrease in the proportion of small RNA. The large molecular weight RNA is derived from the short RNA chains, which are synthesized de novo during the reaction in vitro. The effect of polyanions on the size and nature of the product may be related to structural changes induced in the template rather than to an inhibition of nuclease activity.