The cooperative interaction between multiple protein molecules at specific locations on chromosomal DNA has been a central problem in molecular biology. In this work, we probe for the first time the collision between an active replisome and a stalled or moving RNA polymerase ternary complex using single molecule techniques. In particular, we study the collision between the replicative polymerase from bacteriophage Phi29 and both bacterial and T7 RNA polymerases, representing both true and engineered biological systems. Single molecule tools offer an excellent platform to probe the real time collision between two active protein molecules. Here, we present a novel flow-based DNA stretching assay that allows for simultaneous determination of both the replisome and RNA polymerase locations on individual DNA templates in real time. In this manner, the assay combines mechanical manipulation of DNA templates and fluorescence detection of RNA transcripts. We seek to determine the outcome of DNA polymerase/RNA polymerase interactions, including retention or loss of nascent RNA transcript and stoppage or pausing of DNA replication.