Team:UCSC/Experiments
Experiments
Experiment 0
Description in progress
Figure 1: Creation of pOPPY-UC19-yXXU containing URA3, pUC19 backbone, Lox sites and homologous arms using Gibson Assembly. Transformation of Y. lipolytica to create Y. lipolytica str. LipLox homologous recombination.
Experiment 1
Description in progress
Figure 2: Creation of pOPPY-19-yP via overlap extension PCR of genes involved in progesterone biosynthesis and gibson assembly of this gene cassette with homologous arms and linearized pUC19 backbone. Transformation of Y. lipolytica str. LipLox using homologous recombination to exchange gene cassette. Selection of Y. lipolytica str. PoPPY using 5FOA URA+.
Experiment 2
Description in progress
Figure 3: Creation of pOPPY-XRL2-yP via Yeast Mediated Cloning in S. cerevisiae using linearized pXRL2 and gene fragments. Transformation of Y. lipolytica str. LipLox using pOPPY-XRL2-yP followed by Cre-Lox Recombination and 5FOA URA+ selection to create Y. lipolytica str. PoPPY.
Experiment 3
Description in progress
Figure 4: Yeast Mediated Cloning in Y. lipolytica str. LipLox using linearized pXRL2 and gene fragments. This is followed by cre-recombination of Lip-Lox and 5FOA URA+ selection to form Y. lipolytica str. PoPPY.
Quantification
Description in progress
Figure 5: Progesterone-dependent inactivation of hammerhead ribozyme in the 3’ UTR of GFP mRNA.