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Media, vitamins and so on.

 

 
    About the recalculation of recipes for the arbitrary volume:


    Media.

    LB, LTB

    pH 7.0; p=1.024

     Stock100ml150ml200ml300ml600ml750ml
    1.Bactotriptonesolid1g1.5g2.0g3.0g6.0g7.5g
    2.Bacto Yeast extr.solid0.5g0.75g1.0g1.5g3.0g3.75g
    3.NaClsolid1.0g1.5g2.0g3.0g6.0g7.5g
    4.H2OmQ95.3g143g190.7g286g572g715g
    5.agar topsolid0.65g0.98g1.3g1.95g3.90g4.88g
    6.agar bot.solid1.5g2.3g3.0g4.5g9.0g11.3g
    7.NaOH10N16.7µl25µl33.4µl50µl100µl125µl
    8.MgSO41M1ml1.5ml2.0ml3.0ml6.0ml7.5ml

    LTB: p.1, 3, 4, 7, 8.
    LB: p.1, 2, 3, 4, 7.
    LB/Mg: p.1, 2, 3, 4, 7, 8.

  • it would be better to mix agar after autoclaving. Otherwise it will solidify as heterogeneous substance (much denser on the bottom).
  • LB (low salt)

    For medium with Zeocine pH 7.0

    Stock100ml150ml200ml300ml600ml750ml
    Bactotriptonesolid1g1.5g2.0g3.0g6.0g7.5g
    Bacto Yeast extr.solid0.5g0.75g1.0g1.5g3.,0g3.75g
    NaClsolid0.5g0.75g1.0g1.5g3.,0g3.75g
    H2OmQ95.3g143g190.7g286g572g715g
    NaOH10N16.7µl25µl33.4µl50µl100µl125µl

    M9

    10x

    Conc.Stock50ml75ml100ml
    Na2HPO4x 12 H2O48mM358.14g/M8.60g12.90g17.19g
    KH2PO422mM136.09g/M1.50g2.25g3.00g
    NaCl8.5mM58.44g/M0.25g0.38g0.50g
    NH4Cl18.7mM53/49g/M0.50g0.75g1.00g

    M9:

    Conc.Stock100ml150ml200ml
    M91x10x10ml15ml20ml
    Glucose0.4%10%4ml6ml8ml
    H2O cp.86ml129ml172ml

    add additional components:

    thiamine1mM1M100µl150µl200µl
    MgSO41mM1M100µl150µl200µl
    CaCl20.1mM1M10µl15µl20µl
  • for Petri dishes medium it is necessary to add agar to 3%.
  • NZY

    1 L200ml
    NZ amine10g2g
    NaCl5g1g
    Bacto Yeast extr.5g1g
    MgSO4xH2O2g0.4g
    H2O950ml190ml

    SOC, SOB

    (store at NT).

    Stock200ml400ml
    Bactotriptone 4.0g8.0g
    Bacto Yeast extr. 1.1g2.2g
    NaCl5M0.4ml0.8ml
    KCl1M2.0ml4ml
    H2O ~196ml~391ml
    1. after autoclaving to cool to <60oC;
    2. add
    3. 0.01V 2M Mg2+{= 1M MgCl2 + 1M MgSO4}  
      0.01V 2M glucose;  

    4. 0.2µm filter sterilization;
    5. store at -20oC, (-70oC) in small aliquots.

    SOB - the same solution without glucose.

    Terrific Broth

    pH 7.0

    100ml150ml200ml300ml667ml
    Bactotriptone1.2g1.8g2.4g3.6g8.0g
    Bacto Yeast extr.2.4g3.6g4.8g7.2g16.0g
    Glycerol0.4ml
    0.504g
    0.6ml
    0.757g
    0.8ml
    1.009g
    1.2ml
    1.513g
    2.667ml
    3.362g

    H2O to V90ml135ml180ml270ml600ml
    add after autoclaving:
    KPO4, pH7.0 1.0M10ml15ml20ml30ml66.7ml

    YENB

    pH 7.5: (non-salt medium for electro-competent cells preparation).

    Conc.Stock0.45 L0.5 L0.9 L1.0 L
    Bacto Yeast extr.0.75%solid3.4g3.75g6.75g7.5g
    Nutrient Broth0.8%solid3.6g4.0g7.2g8.0g
    NaOH 10N~243 µl~270 µl~490 µl~540 µl

    2 YT

    %(w/w)200ml1L
    Bactotriptone1.63.2g16g
    Bacto Yeast extr.1.02g10g
    NaCl0.51g5g
    H2O96.9193.8ml969ml

    Other substances.

    IPTG

    C6H18O5S; Mw=238.3g/M;

    Conc.1ml5ml10ml
    IPTG200mg/ml0.2mg1.0g2.0g
  • filter sterilization;
  • store dry powder at 4oC (dry),-20oC (solution in H2O);
  • - for plasmids - per 80mm Petri dish ~4µl,
    - for M13 - per 3ml of top agar ~1-4µl.

    X-Gal

    C14H15BrClNO6 Mw= 408g/M;

    Conc.1ml5ml10ml
    X-Gal20mg/ml (2%)20mg0.1g0.2g
  • store dry powder at 4oC in the dark; solution at -20oC in the dark (in DMFA (or DMSO with the same result)).
  • - for plasmids - per 80mm Petri dish 40µl,
    - for M13 - per 3ml of top agar 40µl.

    Glucose

    C6H12O6 Mw=180.16;

    Conc.Stock5ml10ml50ml100ml150ml200ml
    C6H12O61M180g/M0.90g1.80g9.01g18.02g27.03g36.03g
  • filter sterilization.
  • Maltose.

    100x solution for bacteriophage lambda media;

    Conc.Stock10ml50ml100ml
    Maltose20%solid2g10g20g
  • filter sterilization.
  • Thiamine.

    Vitamin B1 Aneurine; C12H17ClN4OSxHCl; Mw=337.3g/M;

    Conc.Stock1ml5ml10ml25ml
    C12H17ClN4OSxHCl1M337.3g/M0.337g1.69g3.37g8.43g
  • filter sterilization.

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