Simultaneous Analysis of Multiple Promoters: An Application of the PC-GW Binary Vector Series

With the advances in the field of synthetic biology, there is an increasing demand for multi-gene cloning technologies. Molecular cloning to generate multi-gene constructs can be performed by restriction digestion, or by recombination-based cloning strategies such as Gateway(®). This chapter details cloning, transformation, and selection procedures involved in generation of multi-gene expressing transgenic plants. Methods are described for cloning five distinct promoter-reporter fusion constructs into the PC-GW-BAR vector (from the PC-GW vector series) using Gateway(®) technology and meganuclease sites. Further, transformation and selection methods are described for the biofuel crop Camelina sativa from the Brassicaceae family. These methods would be constructive toward generating multi-gene expressing plants for simultaneous expression analysis of five promoters in a short time period.