Rebuilding T7

Summary

T7.1

Wild-type T7 is a superb organism for discovering the primary components of a biological system [Studier, 1972]. However, is the original T7 isolate also best suited for understanding how all parts of the phage are organized to encode a functioning whole? Given our experiences, we decided to attempt to engineer a surrogate genome, which we designated T7.1. Two goals drove our design of T7.1. First, we wanted to insulate and enable independent manipulation of all identified genetic elements. Second, we wanted the T7.1 genome to encode a viable bacteriophage. At the start of this work, we were uncertain how many simultaneous changes the wild-type genome could tolerate.

T7.2

Five goals will drive our design of T7.2; the first four goals revisit or extend those used in the design of T7.1. First, we will specify a genome that only includes elements that webelieve contribute to phage gene expression. Moving beyond our design of T7.1, we will actively erase or delete elements of unknown function. Second, we will specify a genome that does not include any functions that might be encoded via the physical coupling of multiple genetic elements. Third, our design of T7.2 will enable the unique and selection-independent manipulation of each genetic element via restriction enzymes. Fourth, for practical reasons, our design of T7.2 must encode a viable bacteriophage. Fifth, to attempt to make our modeling of gene expression easier, we will use standard synthetic elements in place of the natural elements that regulate transcription and translation. Taken together, our design of T7.2 should specify a genome that is simpler to model andmanipulate, in which we have a putative function for each base pair of DNA involved in phage gene expression. Thus, we hypothesize that T7.2 will also encode a dynamicsystem that is easier to model and interact with, relative to the wild type.