Jun 12, 2019

Public workspaceBG11 medium

DOI
https://dx.doi.org/10.17504/protocols.io.s2pegdn
BG11 medium
  • Roscoff Culture Collection1
  • 1CNRS-Sorbonne Université, Station Biologique, Place G. Tessier 29680 Roscoff FRANCE
  • Roscoff Culture Collection
Daniel Vaulot
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DOI: https://dx.doi.org/10.17504/protocols.io.s2pegdn
Protocol Citation: Roscoff Culture Collection 2019. BG11 medium. protocols.io https://dx.doi.org/10.17504/protocols.io.s2pegdn
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: August 28, 2018
Last Modified: June 12, 2019
Protocol Integer ID: 15151
Keywords: freshwater cyanobacteria, cyanobacteria, bg11 medium medium
Abstract
Medium to grow freshwater cyanobacteria.
Materials
STEP MATERIALS
ReagentBG11 mediumMerck MilliporeSigma (Sigma-Aldrich)Catalog #C3061-500ML
Protocol materials
ReagentBG11 mediumMerck MilliporeSigma (Sigma-Aldrich)Catalog #C3061-500ML
Troubleshooting
Before start
Please refer to our general recommendations to grow cultures :
https://www.protocols.io/private/A48906DC1374AD6281495CB86A8F092F
Prepare using Sigma Aldrich stock
  • Under hood, to 1L of sterile water, add 20 mL of Cyanobacteria BG-11 Freshwater Solution from Sigma
  • Filter the medium on 0,2 microns
ReagentBG11 mediumMerck MilliporeSigma (Sigma-Aldrich)Catalog #C3061-500ML
Prepare from base chemicals
Protocol
NAME
Recipe for standard BG-11 media
CREATED BY
Anna Behle
  • CaCl2 2H2O (3.6 g · L-1)
  • Citric acid (0.6 g · L-1)
  • NaNO3 (149.58 g · L-1)
  • MgSO4 · 7 H2O (7.49 g · L-1)
  • 0.25 M Na2-EDTA, pH 8.0 (0.56 ml · L-1)
For 100x BG11 Stock -N: 
  • Omit NaNO3.
  • 1000x Na2CO3: 20 mg L-1
  • 100x TES-buffer, pH 8.0 (1M), adjust with KOH
  • 1000x K2HPO4 x 3 H2O: 30 mg · mL-1
  • 1000x Fe(III) ammonium citrate (6 mg · L-1)
  • 5000x CuSO4 5 H2O (395 ng · mL-1) (sterilize using a filter)
1000x concentration:
  • H3BO3 (2.86 g · L-1)
  • MnCl2 · 4 H2O (1.81 g · L-1)
  • ZnSO4 · 7 H2O (0.222 g · L-1)
  • Na2MoO4 · 2 H2O (0.390 g · L-1)
  • Co(NO3)2 · 6 H2O (0.049 g · L-1)
For BG11 lacking certain metals (e.g. for working with metal inducible promoters PpetE, PcoaT, PziaA etc., trace metal mix can be prepared lacking these chemicals and used instead of standard trace metal mix.
Fill 1 L bottle with 500 mL ultra pure water. Add stock solutions as shown below.
Stock solutionVolume
100x BG11 Stock10 mL
1000x Na2CO31 mL
1000x K2HPO4 x 3 H2O1 mL
100x TES-buffer10 mL
1000x Trace Metal Mix1 mL
Add ultra pure water to 1 L.
Autoclave.
After autoclaving, add 1 mL 1000x Fe(III) ammonium citrate.
Optional: After autoclaving, add 200 µL 5000x CuSO4
Fill 1 L bottle with 500 mL ultra pure water. Add stock solutions as shown below.
Stock solutionVolume
100x BG11 Stock -N10 mL
1000x Na2CO31 mL
1000x K2HPO4 x 3 H2O1 mL
100x TES-buffer10 mL
1000x Trace Metal Mix1 mL
Add ultra pure water to 1 L.
Autoclave.
After autoclaving, add 1 mL sterile 1000x Fe(III) ammonium citrate.
Optional: After autoclaving, add 200 µL sterile 5000x CuSO4
Fill 500 mL bottle with 250 mL ultra pure water. Add stock solutions as shown below.
Stock solutionVolume
100x BG11 Stock -N10 mL
1000x Na2CO31 mL
1000x K2HPO4 x 3 H2O1 mL
100x TES-buffer, pH = 8.010 mL
1000x Trace Metal Mix1 mL
Add ultra pure water to 500 mL.
Autoclave.
After autoclaving, add 1 mL sterile 1000x Fe(III) ammonium citrate.
Optional: After autoclaving, add 200 µL sterile 5000x CuSO4
Prepare 1.5 % agar: Weigh 4.5 g Bacto Agar. Fill up to 300 mL. Autoclave.
Microwave agar until liquid. Let cool.
In a 50 mL Falcon, add 1 vol 2x BG11 and 1 vol liquid 1.5 % agar. (Note: Usually, one plate requires 30-40 mL total volume.)
When mixture is hand warm, add appropriate antibiotics, if required. Quickly pour plate, avoiding air bubbl