Protocols Commonly Used in the Monroy’s Lab

Northern Arizona University

Training:

       1)  Training part 1
       2)  Training part 2

I) RNA Extraction

        a) Trizol Reagent

        b) RNeasy Protocol

        c) RNeasy protocol - Radha

        d) RNA quantification

        e) RNA isolation -Tissues

         f) Nanodrop procedure

II) cDNA Synthesis

       a) cDNA synthesis Maxima

       b) cDNA synthesis Revert Aid

       c) Agarose Gel Electrophoresis

       d) Gel purification

III) Traditional PCR

        a) PCR reactions

IV) Real Time PCR

        a) qPCR protocol

        b) Set-up BioRad qPCR

        c) Analyzing Data

        d) Making standards

        e) Primer normalization

         f) Data Analysis (Josh)

V) Cell Culture Work

      a) MODE-K cells media

      b) Cell culture

      c) Isolation of Intestinal epithelial cells from mice

      d) Intestinal epithelial Cells primary culture

      e) Mouse myofibroblast purification

      f) Isolation of Lamina Propia (LP) lymphocytes from Mice

      g) Magnetic cell separation

      h) Blood lymphocyte purification

      j) Transepithelial resistance

VI) Toxoplasma Work

        a) Parasite purification

        b) Parasite freezing

VII) Protein work

        a) Indirect fluorescent antibody test (IFAT)

        b) Counterstaining of slides

        c) SDS-PAGE

        d) Western blotting

        e) Chemiluminescence development

        f) Stripping Membrane

        g) Lysis buffer for protein work

        h) Paraffin sections staining

        i) Immunoprecipitation protocol

        j) Kynurenine assay

        k) Mounting media

         j) Western blot - Radha

VIII) Buffers

        a) PBS and buffered formalin

        b) RIPA buffer

IX) Laboratory

        a) Lab book keeping and Expectations