Development of a High-Throughput Physcomitrella patens Transformation System for Biosynthetic Applications

A key challenge for plant synthetic biology is the development of a platform for efficient multi-gene engineering. The moss Physcomitrella patens already has several desirable features for a biosynthetic platform, such as precise genome engineering via homologous recombination, easy upscaling of homogenous cultures, and a fully sequenced genome. However, a lack of methods for rapid and easy multi-gene engineering keeps this system from realizing its potential. Here we seek to fix two major limitations: 1) the lack of a genetic regulatory element toolkit and 2) a transformation process that is bottlenecked by labor. Creating a transgenic moss strain expressing more than one gene product necessitates the use of several promoters and terminators to prevent inappropriate recombination between multiple copies. Thus, new promoters and terminators need to be screened and catalogued. Automation of moss transformation and tissue culture techniques will not only aid our genetic element screen, but will accelerate the advent of a useful and productive moss synthetic biology platform.