ARTIFICIAL SEAWATER BASED AMP1 MEDIUM

LR Moore, A Coe, ER Zinser, MA Saito, MB Sullivan, D Lindell, K Frois-Moniz, J Waterbury, SW

Chisholm (2007).
Culturing the marine cyanobacterium Prochlorococcus.
Limnol. Oceanography: Methods 5: 353-362.

Recipe

Components Final Concentration

Turk’s Island Salt Mix:
NaCl 481 mM
MgSO4·7 H2O 28 mM
MgCl2·6 H2O 27 mM
CaCl2·2 H2O 10 mM
KCl 9 mM
Macronutrients:
NaH2PO4 50 µM
(NH4)2SO4 400 µM
Buffers:
NaHCO3 6 mM
HEPES 1 mM
Trace Metals:
Na2EDTA·2 H2O 0.1170 µM
FeCl3·6 H2O 0.1180 µM
ZnSO4·7 H2O 0.0008 µM
CoCl2·6 H2O 0.0005 µM

NOTE: To avoid contamination, dry nutrients and metals should be weighed

without using spatulas.
PROTOCOL

1. Prepare macronutrient stocks using the highest grade chemicals (i.e.

Sigma Ultra) to avoid trace metal contamination. The chemicals are
dissolved with Type I ultrapure water and filter sterilized through 0.2µm
syringe filter (Acrodisc, Pall # 2006-01) or through sterile vacuum nylon
filtration systems (i.e. VWR # 87006-076) into acid washed sterile
containers in laminar flow hood. Store all macronutrients in separate
containers at 4°C.
Note: The NaH2PO4 solution should be adjusted to pH 7.5 with NaOH
before sterilizing.
2. Prepare trace metals using the highest grade chemicals (i.e. Sigma Ultra)
to avoid trace metal contamination. The trace metal chemicals are
dissolved together with Type I ultrapure water and filter sterilized through
0.2µm syringe filter (Acrodisc, Pall # 2006-01) or through sterile vacuum
nylon filtration systems (i.e. VWR # 87006-076) into acid washed sterile
containers in laminar flow hood and store at 4°C.
3. Prepare buffers using the highest grade chemicals (i.e. Sigma Ultra) to
avoid trace metal contamination. The buffer chemicals are dissolved
together with Type I ultrapure water and filter sterilized through 0.2µm
syringe filter (Acrodisc, Pall # 2006-01) or through sterile vacuum nylon
filtration systems (i.e. VWR # 87006-076) into acid washed sterile
containers in laminar flow hood and store at 4°C.
Note: The HEPES solution should be adjusted to pH 7.5 with NaOH
before sterilizing. NaHCO3 should only be stored for up to one month.
4. Prepare Turk’s Island salts by dissolving each salt in order into Type I
ultrapure water and sterilize by autoclaving.
5. Add filter-sterilized macronutrients, HEPES and NaHCO 3 buffers, and
trace metal mix to sterilized salt mix and store up to one month at room
temperature.