Cultures of 115 SynCom member strains were picked directly from glycerol stocks into 1 ml TSB in 96-well deep-well plates using a 96-well format microplate tip replicator and sealed with breathable plate sealer (AeraSeal, Sigma Aldrich). Cultures were grown at 25°C with 180 RPM agitation for five days and controlled to ensure that a majority of strains grew successfully. Fresh TSB (500 μl per well) was added and cultures were grown overnight, for 12–18 h to harvest metabolically active cells. Cultures were then centrifuged (4000 g, 20 min), washed once with MgCl2, and resuspended in 300μl/well. Cultures were combined, washed, resuspended in MgCl2, and adjusted to OD600 = 0.1. Heat-killed SynCom was prepared by incubating an aliquot of SynCom suspension at 99°C for 30 min. Heat-killed or Live suspensions were used 1000x to inoculate media (final OD600 = 0.0001).
Aeraseal
Manufactured by Merck Group
Sourced in Macao
AeraSeal is a laboratory equipment product designed for sealing and capping a variety of sample containers and microplates. It functions to provide a secure and airtight seal to prevent sample contamination or evaporation.
Automatically generated - may contain errors
Lab products found in correlation
2 protocols using aeraseal
1
Cultivation and Preparation of Synthetic Community
2
High-throughput Butanol Tolerance Screening
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High-throughput phenotyping of the 165-strain Saccharomyces cerevisiae collection was performed in 96-well plates (NUNC, Thermo Scientific, Rockford, IL). Briefly, 10 µl of thawed frozen stock of cells was used to inoculate a 96-well plate containing 190 µl of YPD media. Plates were sealed with breathable tape (AeraSeal, Sigma, St. Louis, MO), covered with a lid and incubated at 30° while shaking for 24 hr, at which time a new subculture was generated by inoculating 190 µl of YPD with 10 µl of the previous culture and grown to log phase for 6 hr. 10 µl of this log-phase culture was inoculated into 190 µl of YPD + 2% butanol, plates were then sealed with an aluminum foil seal (Cryostuff, Vienna, VA, USA) to minimize butanol evaporation, grown with shaking for 24 hr, followed by measurement of the final OD600 using the Tecan M200 Pro microplate reader (Tecan Systems, Inc., San Jose, CA). The average of four biological replicates was calculated to represent tolerance to butanol. Single-strain phenotyping was performed in shaker flasks at 30C under defined media conditions.
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