Articles
THE MOLECULAR BASIS FOR SOMATIC EMBRYO DEVELOPMENT IN CARROT
However, progress has been relative slow and successes have been fragmentary; each laboratory working toward this goal has identified and characterized one or a few such genes.
In recent years, we (and others) have come to understand that part of the impediment to clone isolation from somatic embryos has resided in the strategy behind the library construction and screening which relied almost exclusively on a comparison of transcripts present in callus cells and somatic embryos.
We have taken a new approach to this problem; we prepared a cDNA library from polysomal mRNA of globular staged embryos of carrot, and screened it with a "subtracted cDNA probe" in which globular cDNA was prehybridized with polysomal mRNA from zygotic seedlings to remove all common sequences.
Using this approach, we have been successful in isolating 30 different clones, all of which are enhanced in globular embryos compared to seedlings.
Preliminary sequence analysis on all of these different clones shows that in a single screen, we have isolated clones corresponding to many of the embryo clones isolated by the other labs who have worked in this area over the last seven years, as well as at least 24 new clones which have never been described, and which we are now characterizing.
These results validate the utility of the screen to identify embryo-enhanced genes, and suggest to us that, at the molecular level, callus cells are not very different from the embryos that develop from them.
Indeed, it appears that many callus cells in an embryogenic culture line are actually pre-embryos, and that many of the mRNAs that direct early embryo development are already present.