================================================================================================================== medium no. 30: Gamborgʼs B5 medium, 1 µM 2,4-dichlorophenoxyacetic acid, 0.04 µM kinetin, 0.9% (w/v) agar, pH 5.7 ================================================================================================================== Plant cell lines ================= * :doc:`/cell_lines/rpc00035` Chemicals and stock solutions ============================== (All stock solutions are stored at 4°C) * B5 salt mix Gamborgʼs B5 Medium Salt Mixture, FUJIFILM Wako Pure Chemical Corporation (#399-00621) * Sucrose * B5_VT .. csv-table:: :header: "Chemical", "Concentration (mg/mL)" :widths: auto "Nicotinic acid", "0.4" "Pyridoxine·HCl", "0.4" "Thiamine·HCl", "4" "*myo*-Inositol", "40" * 2,4-D (1 mM) .. csv-table:: :header: "Chemical", "Concentration (mg/mL)" :widths: auto "2,4-D sodium monohydrate", "0.261" (2,4-Dichlorophenoxy)acetic acid sodium salt monohydrate, Sigma-Aldrich (D6679) * Kinetin (1 mM) .. csv-table:: :header: "Chemical", "Concentration (mg/mL)" :widths: auto "Kinetin", "0.215" Dissolve kinetin in small volume of KOH (1 N), and fill up with distilled water * Agar, powder * KOH (1 N) Medium preparation =================== #. Dissolve the following chemicals in approximately 800 mL of distilled water. .. csv-table:: :header: "Chemical", "Amount" :widths: auto "B5 salt mix", "1 bag (1 L)" "Sucrose", "20 g" #. Add following stock solutions, and fill up to approximately 950 mL with distilled water. .. csv-table:: :header: "Stock solution", "Volume (mL)" :widths: auto "B5_VT", "2.5" "2,4-D (1 mM)", "1" "Kinetin (1 mM)", "0.04" #. Adjust the pH of the solution to 5.7 with KOH (1 N), and fill up to 1 L with distilled water. #. Add 9 g of agar to the medium. #. Autoclave the medium at 121°C for 20 min. Formulation ============ .. csv-table:: :header: "Chemical", "Concentration (mg/L)" :widths: auto "KNO\ :sub:`3`", "2500" "(NH\ :sub:`4`\ )2SO\ :sub:`4`", "134" "CaCl\ :sub:`2`\ ·2H\ :sub:`2`\ O", "150" "MgSO\ :sub:`4`\ ·7H\ :sub:`2`\ O", "250" "NaH\ :sub:`2`\ PO\ :sub:`4`\ ·H\ :sub:`2`\ O", "150" "H\ :sub:`3`\ BO\ :sub:`3`", "3" "MnSO\ :sub:`4`\ ·H\ :sub:`2`\ O", "10" "ZnSO\ :sub:`4`\ ·7H\ :sub:`2`\ O", "2" "KI", "0.75" "Na\ :sub:`2`\ MoO\ :sub:`4`\ ·2H\ :sub:`2`\ O", "0.25" "CuSO\ :sub:`4`\ ·5H\ :sub:`2`\ O", "0.025" "CoCl\ :sub:`2`\ ·6H\ :sub:`2`\ O", "0.025" "FeSO\ :sub:`4`\ ·7H\ :sub:`2`\ O", "27.8" "Na\ :sub:`2`\ -EDTA", "37.3" "Nicotinic acid", "1" "Pyridoxine·HCl", "1" "Thiamine·HCl", "10" "*myo*-Inositol", "100" "Sucrose", "20000" "2,4-D sodium monohydrate", "0.261" "Kinetin", "0.0086" "Agar", "8000"