Can vitrified zebrafish blastomeres be used to obtain germ-line chimaeras?

Blastomere cryopreservation plays an important role in maintaining the genetic diversity for valuable fish species. Recently, an original procedure for blastomere vitrification in zebrafish (Danio rerio) was developed in our lab. In the present work, blastomeres from the wild strain embryos, previously vitrified-thawed by this procedure, were injected into embryos from the gold strain in order to assess their ability to colonise the germ-line of recipient embryos. The blastomere survival rate at thawing (higher than 90 percent) as well as the whole number of recovered blastomeres per donor embryo (around 20 percent), were in the ranges previously reported for this vitrification technique. Despite this, only 2 adult chimaeric specimens were finally obtained from a total of 47 injected embryos. Signals of chimaerism were not detected at any stage of development of the chimaeric embryos (somatic chimaerism) or in adulthood (somatic and germ-line chimaerism). In relation to this, difficulties during blastomere insertion are thought to be responsible for the poor results obtained, their aspects being discussed in detail in this work. More improvements to overcome such technical difficulties are needed and, until then, blastomere vitrification may only be of interest for germplasm cryobanking.