Rho is an essential enzyme that is required for the survival of most bacterial species. During transcription, Rho facilitates the process of transcription termination at Rho-dependent terminators across the genome. Using Nascent Elongating Transcript-Sequencing (NET-Seq), the goal of my research project is to determine the specific sites of Rho-dependent transcription termination across the genome of the bacterium Escherichia coli. In addition to the control rpoC::3XFLAG strain, an rpoC::3XFLAG hfq::cat strain was successfully constructed in preparation for NET-Seq experiments.  We expect that the comparison of NET-Seq data for the control strain with the hfq mutant strain untreated or treated with the Rho-inhibitor bicyclomycin (BCM) will allow us to identify sites where Hfq-dependent small RNA (sRNA) modulate Rho-dependent termination across the E. coli genome.  In addition to the control strain and hfq mutant strain, an rpoC::3XFLAG gcvB::kan strain is being constructed to further map sites where the sRNA GcvB modulates Rho-dependent termination across the genome.  Using NET-Seq, the treatment of each strain with or without BCM will allow us to identify the sites of Rho-dependent termination across the E. coli genome in the presence or absence of Hfq or the sRNA GcvB.