How to Cultivate Ectocarpus: Figure 1.

Abstract

This article describes the standard procedure for growing Ectocarpus in the laboratory. The culture is started with partheno-sporophyte (or sporophyte) filaments because this is the stage that is usually maintained in strain collections. The standard medium is Provasoli-enriched natural seawater (PES), but Ectocarpus can also be grown in artificial seawater, which allows more precise control over the culture conditions. The algae can be cultivated either in plastic Petri dishes or in 10-L bottles with bubbling, if large amounts of biomass are required. Standard growth conditions are 13°C with a 12h/12h d/night cycle and 20 µmol photons m(-2) s(-1) irradiance using daylight-type fluorescent tubes. All manipulations of Ectocarpus cultures should be performed in a clean environment (if possible, under a laminar flow hood). Forceps should be dipped in ethanol and allowed to dry under the hood.

Effects of Metal Ions on Stability and Activity of Hyperthermophilic Pyrolysin and Further Stabilization of This Enzyme by Modification of a Ca 2+ -Binding Site

“…One unit (U) of activity was defined as the amount of enzyme required to increase the A 335 value by 0.01 per min. To determine the activity of the enzyme in artificial seawater (ASW), 20 l of enzyme sample in buffer A was diluted with 180 l of ASW (450 mM NaCl, 10 mM KCl, 9 mM CaCl 2 , 30 mM MgCl 2 , 16 mM MgSO 4 , pH 7.8) (28) and then mixed with 200 l of 0.5% (wt/vol) azocasein in ASW. The samples were then assayed for enzymatic activity as described above.…”

DISTAG/TBCCd1 Is Required for Basal Cell Fate Determination in Ectocarpus

“…Strain cultivation, genetic crosses, raising of sporophytes from zygotes, and isolation of meiotic families were performed as described previously (Coelho et al, 2012a(Coelho et al, , 2012d). Gametes of Ectocarpus are able to develop parthenogenically to produce haploid partheno-sporophytes, which are identical morphologically to the sporophytes that develop from diploid zygotes (Peters et al, 2008;Coelho et al, 2011).…”

“…After dissection, the material was kept at 13°C under standard culture conditions (Coelho et al, 2012e). Wild-type and dis sporophyte apical filament cells were isolated by microdissection 7 d after the emergence of the first upright filaments (20 d after gamete release); gametophyte apical filament cells were isolated by microdissection 15 d after meiospore release.…”

The Ectocarpus IMMEDIATE UPRIGHT gene encodes a member of a novel family of cysteine-rich proteins with an unusual distribution across the eukaryotes

“…The wild-type individuals used in this study were strain Ec32 (CCAP1310/4), a brother of the original imm mutant strain Ec137. Ectocarpus was cultivated as described previously (Coelho et al, 2012b).…”

“…strain Ec32 gametes using the transfection reagent HiPerFect (Qiagen). One microlitre each of 0.5 µg/µl solutions of HPLC purified siRNAs in 1× Universal siMAX siRNA Buffer (MWG Eurofins) was mixed with 12 µl HiPerFect transfection reagent in a final volume of 100 µl of natural seawater, vortexed to mix and incubated for 10 min at room temperature before being added dropwise to 100 µl freshly released gametes (Coelho et al, 2012b) in natural seawater in a Petri dish. After rotating gently to mix, the Petri dish was incubated overnight at 13°C.…”