Y2H growth curve assay
S. cerevisiae strain PJ69-4A, was transformed containing plasmids
with full length PvPGIP1 or PvPGIP2, and strain PJ69-4α was transformed
one of the PGs (AnPG2, BcPG1, BcPG2, FmPG3). The yeast transformations
were carried out using standard protocols (Truong & Gietz, 2007). 500µL
of overnight stock cultures of these samples were grown at 30°C with
orbital shaking at 250 RPM in -T or -L YNB respectively with 5%
dextrose. After 16 hours, 10µL of each PGIP and PG containing yeast was
mated together in YPD for 24 hours. A 1/60 dilution of each culture was
made into -LT for approximately 48 hours. Finally, a 1/300 dilution of
the mated yeast in the -LT was then made into 3mL of -HTL YNB. Each
sample had 3 biological replicates. A plate reader was used to measure
the OD at 600nm every 24 hours for 4 days. The average OD across the 3
biological replicates were then plotted. The same procedure was applied
for the truncated versions of the PvPGIPs. Data was collected using the
Gen5TM Data Analysis Software (BioTek Instruments).