Team:UNIK Copenhagen/Green Lab
Green Lab
We will make a stable and transient transformation of moss, Psychometrilla patens. We will make two linear gene constructs for stable transformation in moss. One construct is going to contain a gene encoding an antifreeze protein and the other construct will contain a stilbene synthase-gene (STS), making the final enzyme in the resveratrol pathway. Both constructs contain regions homologous to the moss genome, so they will integrate via homologous recombination in moss. The constructs furthermore contains the ZmUbi-promoter, resistance gene and YFP, to confirm transformation. We will also make a transient moss transformation with a vector containing the ZmUbi-promoter followed by YFP, to confirm the function of the promoter in moss.
Due to time constraints, we will also transform tobacco and E.coli. We will transform tobacco with the STS-gene, using USER-cloning and Agrobacterium tumefaciens transformation, in order to quickly confirm the functionality of the gene (moss takes a long time to grow!) by measuring the presence of resveratrol with Liquid chromatography-mass spectrometry. Furthermore we will transform E. coli with the antifreeze gene which originate from the spruce budworm, to quickly see whether the gene has an effect on temperature sensitivity. We will perform experiments at low temperatures to measure the growth of the bacteria when exposed to cold. We will also make a his-tag version of the antrifreezeprotein, to confirm the presence of the protein by western blotting.
one of our petri dishes with moss
Moss growing facility