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Protocols Fission Yeast Biochemistry Molecular Biology Tissue Culture/Cell Biology Bioinformatics Others Fission Yeast Nurse lab manual Expressing protein from pRep transformed yeast Preparation of cell lysate for SDS-PAGE (S. Wu) Isolation of diploids (general) (R. Geyer) Isolation of diploids (specific) (R. Geyer) Genomic C-terminal tagging by PCR (C. Zhou) Fractionation of cytoplasmic and nuclear proteins (V. Seibert) FACS (V. Seibert) Immunofluorescence by formaldehyde fixation (E. Rhee) Antibiotic sensitivity (E. Rhee) Preparation of genomic DNA for PCR Determining protein stability in vivo by cycloheximide chase (S. Wee) Freezing S. pombe cells (S. Wee) Calcofluor staining (F. Arslan) Preparation of S. pombe lysate for LC-MS/MS analysis (S. Wu) Large scale S. pombe cell lysate preparation for biochemistry - Coffee grinder method (E. Amoah-Buahin) Making haploids from Bioneer h+/h+ diploids (S. Wu) Polysome fractionation and mRNA extraction (P. McQuary) Biochemistry Bacterial Expression in BL21 Flex column assembly and use Concentrating protein Affinity Purification of antibodies (V. Seibert) Purification of monoclonal antibody from hybridoma supernatant (C. Zhou) Gradient minigels Large gradient gels (S. Wee) Vorum silver staining protocol (compatible with MALDI MS) Fractionation of cytoplasmic and nuclear S. pombe proteins In-gel digestion of proteins for MALDI MS Multidimensional 2D gel analysis of S. pombe lysate Affinity purification of protein complexes with IgG/proA (Version 1 - R. Geyer) Affinity purification of protein complexes with IgG/proA (Version 2 - Optimization by Z. Sha) TEV cleavage and TCA precipitation In vitro ubiquitination assay with IPed cullin complexes (M. Schmidt) Autoinduction of fusion proteins in BL21 (E. Rico-Bautista) Tris-Tricine gels for small proteins (E. Rico-Bautista) Preparation of S. pombe lysate for LC-MS/MS analysis (S. Wu) Affinity purification of GFP fusion proteins from S. pombe or mammalian cells (J. Scheliga) Polysome fractionation and mRNA extraction (P. McQuary) FLAG purifications from transfected mammalian cells (J. Scheliga) Polysome profiling with mammalina cells (DAW) Molecular Biology Competent bacteria Miniprep RNA preparation and RT-PCR (S. Wee) Northern Blotting Yeast 2H screening (A. Piau) Tissue Culture/Cell Biology Purification of monoclonal antibody from hybridoma supernatant Production of baculoviruses Expression analysis of recombinant proteins Production of adenoviruses (H. Schulz) Amplification of adenoviruses (C.C. Yang) Immunofluorescence staining of LNCaP prostate cancer cells (E. Rico-Bautista) FACS (D.A. Wolf) High efficiency transfection of LNCaP and PC-3 cells (C.C. Yang) Bioinformatics Assembling gene lists from InterPro protein domains (also includes gene name conversion with Panther) - (C.-C. Yang) Others Sporulation of budding yeast (S. Wee) Transformation of budding yeast (S. Wee)