Based on _BFG-Y2H_reagents10262013.docx from Nozomu Yachie (Modified from Haiyuan Yu’s protocol).

1. Common solutions

40% glucose

Dissolve 400g of glucose in 1L of ddH2O, filter to sterilize. Use 50mL for 1L medium.  



50% glycerol

  Add 1000mL glycerol to 1000mL dH2O (2L in total) and autoclave.                        



2. Amino acid solutions

12mg/mL Adenine solution

  6.00g of Adenine sulfate (hemisulfate) in 500mL of ddH2O. Autoclave and filter to sterilize. Use 15mL for 1L medium. After a while you make the adenine sulfate, it precipitates. Don't worry about it, keep using it after swirling a bit. Alternatively, you can heat it up (75C for 30min) and it dissolves again.  



100mM Histidine

  7.76g of Histidine in 500mL total volume of ddH2O. Autoclave and filter to sterilize and wrap the bottle in foil. Use 8mL for 1L medium to supplement 1X His, but some media in this protocol require 80mL for 1L medium to supplement 10X His.                         



100mM Leucine

6.56g of Leucine in 500mL total volume of ddH2O. Autoclave and filter to sterilize. Use 8mL for 1L medium.                                                                                       



40mM Tryptophan

4.08g of Tryptophan in 500mL total volume of ddH2O. Autoclave and filter to sterilize, wrap the bottle in foil and keep at 4C°. Use 8mL for 1L medium.                                                                                        




20mM Uracil

1.21g of Uracil in 500mL total volume of ddH2O. Autoclave and filter to sterilize. Note that it may be necessary to heat the water to get the uracil into solution. Use 8mL for 1L medium.                                                                                        



3. Drugs

10mg/mL Cycloheximide solution

  100mg to 10mL of pure EtOH. This gives you 10,000X conc. Use 100uL for 1L medium.                                                                                                                                            



1000x Doxicycline solution

Working (final) concentration for BFG-Y2H is 10ug/mL. Add 1000 mg of Doxycline to 100 ml of distilled water. Filter, aliquot 10mL and store at -20 degree. Do not thaw and freeze more than 5 times. Use 1mL for 1L medium.  



1M 3-aminotriazole (3-AT)

Dissolve 840 mg of 3-AT in 10 mL Milli-Q H2O. Filter sterilize. Use 1mL per liter.                                                                                                                                           

Dan Yamamoto-Evans 2019/11/29 18:53

  • interactome/reagent_stock.txt
  • Last modified: 2019/11/29 19:10
  • by dan