From cells to embryos to rooted plantlets in a mist bioreactor
A mist bioreactor using a disposable bag as culture chamber was used to propagate carrot embryogenic cells into rooted plantlets. The best operating configuration was akin to a vertical hanging garden using 50 90 mm nylon mesh for explant attachment. Cells spray inoculated into the reactor were 51.2 viable. Misting cycle and aeration conditions were studied and showed that under the same hourly volumetric nutrient feed and 0 VVM, embryo development in the reactor was best using a 0.3 min on/2.7 min off misting cycle, yielding about 23 post heart stage embryos. Compared to 0 VVM, 3 CO2 enrichment improved embryo development in reactor culture. Spray inoculated cells also attached to several vertically hung poly l lysine coated strips and then developed in situ into embryos. Cell attachment was significantly improved when they were suspended in salt free sucrose solution during spray inoculation. Almost 90 of the originally attached cells remained on the nylon mesh 24 h later after spraying with B5 medium in the mist reactor. Strip grown embryos had the same post heart stage ratio but shorter overall length compared to those developed on a horizontal platform. Young plantlets developed uniformly up and down the hanging strips and did not detach after 3 weeks of culture suggesting this technology may prove useful for improving micropropagation.
Plant Cell, Tissue and Organ Culture (PCTOC)