tobacco BY-2 - (May/30/2008 )

does anyone know where I can find a protocol to make BY-2 cells?

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if you do not mind having a ‘non-clonal’ cell line, you can start them quickly by surface sterilizing BY2 seeds and germinating on B5 root media (or any media without plant hormones) with sucrose (about 50mL in a 125mL shake flask). After 14-21 days on a gyratory shaker, replace the media with an MS-based media with hormones. Trick is to keep the cell concentration high initially. You may need to let the cells settle and pipette off some of the material on top and replace with fresh media after a few weeks to accumulate more suspended cells. Once the cell density is high enough (3-6 weeks, maybe longer) you should have a suspension that is culturable. Subculture 20% v/v every 2-3 weeks for a few culture periods until they look healthy and the large aggregates start to disperse… then you can start to subculture them with fewer cells on the desired time frame.

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There.. Late and transient, but there is always a helping hand here!

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