Worm Breeder's Gazette 13(2): 66 (February 1, 1994)
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.
As part of the Genetic Toolkit Project funded by the National Institutes of Health, we have generated a number of recessive lethal mutations on the left arm of chromosome Ill in the nematode Caenorhabditis elegans. Three gamma induced mutations and 110 EMS induced mutations were generated in the strain BC4121 which) has the genotype sDp3 (lll,f); dpy-17 ( e164 ) unc-32 ( el89 )/ dpy-17 ( e164 )lin- l2 ( n941 ).Balanced mutations were recovered only on the dpy-17 unc-32 chromosome and maintained in Unc-32 worms of the genotype sDp3 ; dpy-17 (let-x) unc-32 / dpy-17 (let-x) unc-32 .
The free duplication sDp3 covers 95% of the left arm of chromosome III and is not known to undergo recombination with the normal homologues. As a genetic balancer, sDp3 allows the precise definition of the region in which mutations are generated as well as permitting the maintenance of lethal carrying mutants as chromosomal homozygotes. sDp3 covers dpy-17 but not unc-32 ,hence we are able to distinguish homozygous dpy-17 unc-32 animals which carry the duplication from those which do not. Thus, recessive lethal mutations can be found by looking for the absence of fertile Dpy-17 Unc-32 animals.
We have recombinationally mapped the lethal mutations and found that they all lie between 8.4 mu to the left and 2.5 mu to the right of dpy-17 .Seventeen of the mutations are inseparable from dpy-17 ,and 59 lethals map between dpy-17 and unc-32 .Twenty of the 29 mutations mapping to the left of dpy-17 are within 1 mu of this marker.
A rough analysis of the blocking stages which result from these mutations was performed, and it was found that 10% of the 113 mutations result in worms dying prior to hatching. Forty-one percent of the mutations result in developmental arrest during early larval stages (L1/L2), 16% arrest development between mid and late larval stages, 21% result in sterile adult individuals and one mutation has a maternal lethal effect. Although these blocking stages are based only on approximate size of developmentally blocked worms after seven days, it seems apparent that there is a bias toward genes necessary early in development in this region of the genome.
lnter se complementation testing has so far revealed that these mutations represent at least 48 different complementation groups ( let-701 to let-748 ).and at least 2 deficiencies (sDf 120 and 121). While we would not expect to pick up large deficiencies using sDp3 due to gene dosage problems, these results indicate that it is possible to pick up small deficiencies. We are planning a larger, similar mutation screen using U.V. radiation to generate more deletions in the same area. These will assist in the further mapping of mutations on linkage group III (Ieft).
In order to correlate these lethal mutations with the physical map, we are presently using cosmid injection to produce transformed strains which carry cosmids constructed from linkage group III (left) DNA (obtained from the St. Louis genome sequencing group). We plan to perform complementation tests with these transformed strains and thereby map a number of our lethal mutations to cosmids. As this region is presently being sequenced by the Genome Sequencing Project is expected that most, if not all, of our lethals will be mapped to cosmids already sequenced. As there are approximately 125 cosmids spanning the region in which the lethal mutations in this screen were generated, we expect to find almost one mutation per cosmid on average. This large number of genetic markers will facilitate a high resolution correlation between the genetic and physical maps, and reveal information regarding the function of the coding regions exposed during sequencing.
We are also in the process of testing these mutations against previously named genes, including those generated by Bucher and Greenwald (1). It is expected that a small number of our lethals will be allelic to previously published genes, at which time we will rename the lethals appropriately and contact the people involved. If anyone has mutations to which our lethals may be allelic and would like to be notified of further developments, please contact David Baillie by mail or e-mail at
dbailliee@darwin.mbb sfu ca
(1) BUCHER, ELIZABETH A. AND IVA GREENWALD. 1991. A Genetic Mosaic Screen of Essential Zygotic Genes in
Caenorhabditis elegans. Genetics 128, 281-292