Journal: Microbial Cell Factories

Article Title: Fate of the UPR marker protein Kar2/Bip and autophagic processes in fed-batch cultures of secretory insulin precursor producing Pichia pastoris

doi: 10.1186/s12934-018-0970-3

Figure Lengend Snippet: Post-induction level of intra- and extracellular Kar2/Bip in controlled bioreactor fed-batch culture. Cell growth and IP production were carried out in controlled bioreactor fed-batch culture as described previously . a Concentrations of glycerol (open triangles), biomass (optical density: open squares), IP (filled circles), and methanol (solid line) are given. The dashed vertical line indicates the end of the glycerol batch (pre-induction) and the start of the methanol feeding phase (post-induction). b Immunoblot probing for the time-dependent change of intra- and extracellular Kar2/Bip in post-induction samples. c Extracellular proteins detected by Coomassie blue staining of SDS-PAGE gel indicating proteins identified by mass spectrometry. The positions of full-length Kar2/Bip (~ 74 kDa, filled arrow) and the truncated approximately centrally cleaved Kar2/Bip fragment (~ 35 kDa, open arrow) are indicated. b , c M denotes the molecular weight marker and the numbers on the bottom of the lanes the post-induction sampling time points. Please note the IP (7 kDa) is not detectable on conventional SDS-PAGE gels (see also ). Please also note that immunoblot and SDS-PAGE gel are aligned for the same molecular weight scale

Article Snippet: Samples (loading volume 7 μL) were run on 12% SDS-PAGE gels prior to Coomassie Brilliant Blue staining or electroblotting onto PVDF membranes (Bio-Rad, Hercules, USA) at 15 V for 45 min.

Techniques: Western Blot, Staining, SDS Page, Mass Spectrometry, Molecular Weight, Marker, Sampling