Worm Breeder's Gazette 14(5): 20 (February 1, 1997)
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
Dept. of Molecular Biology and Genetics Johns Hopkins University School of Medicine 725 N. Wolfe St./515 PCTB Baltimore, MD 21205
Chromosomally-integrated transgenes that express GFP throughout
embryogenesis could serve as useful chromosomal markers, enabling worm
breeders to genotype embryos, when the use of larval/adult markers is
not possible. We have generated such a GFP transgene (pJH1.16) using
the pes-10 promoter. Embryonic expression of pJH1.16 is detected as
early as the 28-cell stage in all somatic blastomeres; by the 2-fold
stage, expression becomes restricted to the gut lineage, and persists
there for the remainder of development. We have generated an integrated
array containing this transgene using dpy-20 DNA as a selectable marker.
By chance, this array integrated on the X chromosome; the resulting
strain was outcrossed six times (JH103) and has a mild Dpy phenotype.
We realized that this X-linked transgene could be used to
determine the chromosomal sex of embryos and young worms before the
appearance of any obvious sexual dimorphism. To test this prediction,
we mated JH103 males to tra-2(q122) (spermless) hermaphrodites and
scored resulting embryos for GFP expression. Green and non-green
embryos were transferred to separate plates and allowed to grow to
adulthood to score their sex. Since hermaphrodites receive their X
chromosomes from both parents, whereas males receive their single X
chromosome from their mothers, we expected all green embryos to be
hermaphrodites, and all non-green embryos to be males. Results were as
follows:
Stage at which GFP Green Non-green
expression was scored
28-cell to comma 7/7 hermaphrodites 9/9 males
comma to 2 fold 3/3 hermaphrodites 3/3 males
3-fold 3/3 hermaphrodites 1/3 males
L1 10/10 hermaphrodites 7/7 males
TOTAL 23/23 hermaphrodites 20/22 males
In summary, 43 out of 45 embryos/L1s were sexed correctly. The
2 embryos which were sexed incorrectly were both 3-fold, a stage at
which gut GFP expression is sometimes harder to detect. We conclude
that this strain can be used reliably to sex embryos, with the possible
exception of 3-fold embryos.
The availability of a transgene that expresses reliably
throughout embryogenesis should prove to be a useful tool for tracking
the segregation of specific chromosomes, and has many conceivable uses
besides the specific one described here. Transgene pJH1.16 and strain
JH103 are available to anyone interested. (Contact Matt at
mwallenf@welchlink.welch.jhu.edu).
We are grateful to Andy Fire for creating the GFP used in this study.