Worm Breeder's Gazette 9(3): 23

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Transposon Tagging of unc-44 and dpy (rh1007)

V.I. Wheaton, A.J. Otsuka and E.M. Hedgecock

Figure 1

In the last Gazette (Vol. 9, No. 2), we reported putative Tc1 
mutations in a novel dumpy gene, dpy (rh1007), and in unc-44,an axonal 
outgrowth gene. The analysis of these genes should yield insights into 
the molecular interactions underlying morphological and neural 
Southern blots of the dpy (rh1007) DNA probed with Tc1 reveal only 
one obvious band (at 2.7kb for Hind III digests) above those observed 
both in the Bristol strain and in a spontaneous revertant. This result 
indicates the initial insertion event took place either in the Bristol 
chromosome or in a region of Bergerac chromosome devoid of Tc1 
elements. To further demonstrate the association of the mutant 
phenotype with the unique Tc1 insertion. DNA from strains produced by 
three factor crosses with flanking markers (a dpy-10 deficiency mnDf31 
on the left and unc-4 (e120) on the right, see figure) has been 
isolated and subjected to Southern analysis. Recombination to the left 
of rh1007 should produce nondumpy, uncoordinated offspring which lack 
the unique 2.7kb Hind III fragment. Conversely, progeny which are 
dumpy and uncoordinated should only arise from a crossover event 
between rh1007 and unc-4 and should retain the 2.7kb fragment. Six 
recombinants thus far compared fit the expected pattern. The three of 
these which are non-dumpy and uncoordinated strains lack the 2.7kb 
fragment, whereas the three which are both dumpy and uncoordinated 
retain this fragment upon Southern analysis. Therefore, the dumpy 
phenotype co-segregates with the Tc1-containing 2.7kb restriction 
fragment. Mutant worm DNA migrating at this molecular weight on 
agarose gels is currently being isolated and cloned into the lambda 
vector Charon 30. 
{Figure 1}
In the case of unc-44, the six or more extra bands associated with 
the mutation suggest that the initial insertion event which generated 
the uncoordinated phenotype took place in the Bergerac chromosome. 
Mapping the adjacent transposons is required in order to determine 
which of the many Tc1 elements produces the mutant phenotype. Toward 
this end, recombinant strains have been constructed from two markers 
close to the unc-44 locus (bli-6 and unc-5), and DNA is being isolated 
from them for Southern analysis.

Figure 1