国产91亚洲福利精品一区二区,国产综合成人久久大片91,国产成人精品久久综合,久久久91精品国产一区二区三区,91福利国产在线在线播放,91精品国产高清久久久久久91,91精品国产福利在线观看麻豆,国产免费一区二区三区在线观看

Harvest and Preservation 

1. Mix the components in the order listed. When the medium is added to the DMSO the solution will warm up due to chemical heat.
2. Harvest cells from a culture that is at or near peak density by filtration and centrifugation at 800 x g for 5 min.
3. Adjust the concentration of cells to at least 2 x 10⁶/mL in fresh medium.
4. Mix the cell preparation and the cryoprotective solution in equal portions.
5. Dispense in 0.5 mL aliquots into 1.0 - 2.0 mL sterile plastic screw-capped cryules (special plastic vials for cryopreservation).
6. Place vials in a controlled rate freezing unit. From room temperature cool at -1°C/min to -40°C. If freezing unit can compensate for the heat of fusion, maintain rate at -1 C/min through heat of fusion. At -40°C plunge ampules into liquid nitrogen. Alternatively, place the vials in a Nalgene 1°C freezing apparatus.  Place the apparatus at -80°C for 1.5 to 2 hours and then plunge ampules into liquid nitrogen.  (The cooling rate in this apparatus is approximately -1°C/min.)  
7. Ampules are stored in either the vapor or liquid phase of a nitrogen refrigerator.
8. To establish a culture from the frozen state place the vial in a 35°C water bath.  Immerse the vial to a level just above the surface of the frozen material. Do not agitate the vial.   Immediately after thawing, do not leave in water bath, aseptically remove the contents of the ampule and inoculate into a T-25 tissue culture flask containing 10 mL of uninoculated ATCC medium 1525.
9. Incubate at 25°C with the cap screwed on tightly.
10. Once the culture is established, vigorously agitate the flask and aseptically transfer 0.5 mL to 10.0 mL of uninoculated ATCC medium 1525.
11. Follow the protocol for maintenance of culture.

Carr M, et al. Molecular phylogeny of choanoflagellates, the sister group to Metazoa. Proc. Natl. Acad. Sci. USA. 105: 16641-16646, 2008. PubMed: 18922774

Dayel MJ, et al. Cell differentiation and morphogenesis in the colony-forming choanoflagellate Salpingoeca rosetta. Dev. Biol. 357: 73-82, 2011. PubMed: 21699890

Salpingoeca sp. ATCC 50818 genome sequencing project Available from: NCBI BioProject