13 Prochordata
149
11111 Comments
Fig. 5. Embryonic cells of the colonial protochordate Botryllus
schlosseri in 4-month-old primary
culture. x 225. From Rinkevich, B.
and Rabinowitz, C., 1994, with permission
An alternative for the dissociation of embryos is chemical treatment. For
this:
- Following the last rinse with WS, wash again with Ca 2 + - and Mg2+_
free saline
- Treat the embryos with I ml trypsin (0.25%)/EDTA (0.02%) for 35 min at room temperature
- Centrifuge the dissociated cells, and suspend in BCCM.
11111 Results
In the culture, individual cells, cellular aggregates, and fragments of embryos may be present together. The aggregates are easily dispersed. The
culture attains confluency after 5 weeks, at which time subculture becomes possible (Fig. 5). Rinkevich and Rabinowitz (1994) have obtained
several continuous cell lines by repeating subculture.
Subprotocol 4
Palleal Buds of Colonial Protochordates (Tunicata)
11111 Materials
- Dissecting microscope
- Centrifuge and centrifuge tubes
- Dissecting tray
- Homogenizer
- Scissors
- Forceps
- Pasteur pipettes
- Sterile syringe with a 28 G needle
- Eppendorf vials
- I5-ml plastic tubes
- 24-Well plates
Equipment