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From 2008.igem.org

Contents 1 LB agar 1.1 Materials 1.2 Procedure 2 LB + amp100 2.1 Materials 2.2 Procedure 3 LB+kan20+amp50 3.1 Materials 3.2 Procedure 4 LB+sp100 4.1 Materials 4.2 Procedure 5 BG-11 agar 5.1 Materials 5.2 Procedure 6 BG-11 + 5% LB 6.1 Materials 6.2 Procedure 7 BG-11+sp2.5+sm2.5 7.1 Materials 7.2 Procedure

LB agar

Materials

• 25 g LB broth mixture (Difco)
• 15 g agar
• 1 L ddH₂O

Procedure

1. Mix dry ingredients together with 1 L ddH₂O using a magnetic stir bar.
2. Autoclave to sterilize. Let cool to 55C.
3. Pour 28 ml LB solution for thin plates; 50 ml for thick plates.

LB + amp₁₀₀

Materials

• 1 L LB agar
• 1 ml 1000X ampicillin stock solution

Procedure

1. Make LB agar as directed.
2. When mixture is cooled to 55C, add 1 ml 1000X ampicilllin stock solution.
3. Plate as usual.

LB+kan₂₀+amp₅₀

Materials

• 1 L LB agar
• 0.5 mL 1000X ampicillin stock solution
• 0.4 mL 1000X kanamycin stock solution

Procedure

1. Make LB as directed.
2. After cooling to 55C, add ampicillin and kanamycin.
3. Plate as usual.

LB+sp₁₀₀

Materials

• 1 L LB agar
• 1.0 mL 1000X spectinomycin stock solution

Procedure

1. Make LB as directed.
2. Cool to 55C, then add spectinomycin.
3. Plate as usual.

BG-11 agar

Materials

• 940 ml nanopure H₂O
• 20 ml 50X thiosulfate (final conc. of 1mM)
• 20 ml 50X TES buffer, pH 8.0 (final conc. of 10mM)
• 20 ml 50X BG-11
• 15 g agar

Procedure

1. Combine agar and nanopure H₂O. Stir mixture with a magnetic stir bar.
2. Autoclave to sterilize. Cool to 55C.
3. Add TES, thiosulfate, and BG-11
4. Test pH (should be ~8.0).
5. Plate 28 ml for thin plates; 50 mL for thick plates.

BG-11 + 5% LB

Materials

• 1.25 g LB broth mixture (Difco)
• 15 g agar
• 940 ml nanopure H₂O
• 20 ml 50X BG-11
• 20 ml 50X thiosulfate
• 20 ml 50X TES buffer

Procedure

1. Combine LB and agar with the nanopure H₂O.
2. Autoclave to sterilize.
3. Add TES, thiosulfate, and BG-11.
4. Test pH (~8.0).
5. Plate as usual.

BG-11+sp_2.5+sm_2.5

Materials

• BG-11 agar
• 0.15 ml 1000X spectinomycin stock solution
• 0.3 ml 1000X streptomycin stock solution

Procedure

1. Make BG-11 agar as directed.
2. After adding TES, thiosulfate, and BG-11, add spectinomycin and streptomycin.
3. Plate as usual.

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