Small molecule stocks for TC

Antibiotic concentrations

Reference: Selection antibiotics (ThermoFisher)

Name

Temperature

Stock conc

Common working conc.

293T conc. KL 2023.12.11

Blasticidin

-20°C

10 mg/mL (1kx)

1 - 20 µg/mL

10 µg/mL (1:1000) by 4-5 dpt

Geneticin (G-418)

-20°C

? mg/mL (1kx)

200 - 500 µg/mL

400 µg/mL (1:100) - no death observed

Hygromycin

-20°C

50 mg/mL (250x)

200 - 500 µg/mL

200 µg/mL (1:250) by 6-8 dpt

Puromycin

-20°C

1-10 mg/mL (1-10kx)

0.1 - 10 µg/mL

1 µg/mL (1:10,000) by 3-4 dpt

Zeocin

-20°C

100 mg/mL (100x)

50 - 400 µg/mL

1,000 µg/mL (1:100) >8 dpt

Other stock and working concentrations

Name

Temperature

Stock conc

Common working conc.

Doxycycline

-20°C

1 mg/mL (1kx)

1 ng/mL - 1 µg/mL

Grazoprevir (GZV)

-80°C

1 mM (1kx)

1 nM - 1 µM

Polybrene (Hexadimethrine bromide)

-20°C

5 mg/mL (1kx)

5 - 8 µg/mL

Indole-3-acetic acid (Auxin, IAA)

  1. Resuspend IAA in ethanol at a ratio of approximately 11 uL ethanol to 2 mg IAA. Vortex until dissolved.

  2. Add mixture to ELGA water to achieve a final IAA concentration of 1.75 mg/mL (10mM).

  3. Sterilize with 0.22 µm filter.

  4. Separate into 50 uL aliquots and store at -20°C.

Indole-3-acetamide (Auxin precursor, IAM)

  1. Resuspend IAM in ethanol at a ratio of approximately 20 uL ethanol to 1 mg IAM.

  2. Add mixture to ELGA water to achieve a final IAM concentration of 1.75 mg/mL (10mM).

  3. Sterilize with 0.22 µm filter.

  4. Separate into 50 uL aliquots and store at -20°C.

50mM O6-Benzylguanine (O6BG)

  1. Dissolve 12 mg of O6BG in 1ml of DMSO.

  2. Sterilize with 0.22 µm filter.

  3. Separate into 50 uL aliquots and store at -20°C.

25mM Guanine

  1. Dissolve 7.5 mg of guanine in 2 ml of 0.2N NaOH/Water. (Guanine can not be dissolved with DMSO/Water)

  2. Sterilize with 0.22 µm filter.

  3. Separate into 50 uL aliquots and store at -20°C.

50 μM AP1903/Rimiducid (1000x stock)

  1. Dissolve 1 mg of AP1903 in 14.2 mL of .1 % DMSO/water.

  2. Sterilize with 0.22 µm filter.

  3. Separate into 50 uL aliquots and store at -20°C.

Puromycin, 1 mg / mL (1,000x stock)

We use dissolved 10 mg/mL puromycin (Invivogen, cat. ant-pr-1). It comes as 10 tubes x 1 ml @ 10 mg/mL dissolved in HEPES buffer (5 mM, pH 7.3).

To dissolve a single 1 mL @ 10 mg/mL tube:

  1. Make 5 mM HEPES from the 1M HEPES stock: 45 µL of 1M HEPES + 9 mL sterile water

  2. Add 1 mL of 10 mg/mL puro to the 9 mL of 5 mM HEPES

  3. Sterilize with 0.22 μM filer

  4. Split into 50 μL aliquots and store at -20°C