Students often memorize the definition of a transcriptional promoter but fail to fully understand the critical role promoters play in gene expression.  This laboratory lesson allows students to conduct original research by characterizing functional regions within known prokaryotic promoters.  Students begin the lesson by learning the properties of transcriptional promoter DNA sequences.  They design mutations for a constitutive promoter and discuss their designs as a class to choose which mutations to clone and characterize.  This lesson provides an easy way for faculty with limited time and budgets to give their students access to real research in the context of traditional teaching labs that meet once a week for under three hours.  The pClone Red Genetics lesson uses synthetic biology methods and makes cloning so simple that we have 100% success rates with sophomores taking Genetics.  Students archive promoter sequences and their performances under standard conditions.  The database permits synthetic biology researchers around the world to find a promoter that suits their needs and compare relative levels of transcription.  The core methodology in this lesson is identical to the core methodology in the companion Introductory Biology Lesson by Campbell and Eckdahl. The methods are reproduced in both lessons for the benefit of readers.  The two CourseSource lessons provide the detailed information needed to reproduce the pedagogical research results published in CBE – Life Sciences Education by Campbell et al., 2014.