Worm Breeder's Gazette 11(5): 19

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Cloning daf-10 and osm-1

Steven Stone and Jocelyn Shaw

We are interested in studying the molecular genetics of sensory 
neuron development.  Two genes previously implicated in sensilla 
development, particularly in the development of sensory cilia, are daf-
10(IV) and osm-1(X) (Perkins et.  al.  1986 Dev.  Biol.  117, 456487). 
Mutant animals have a variety of ciliary defects.  However, the major 
defect appears to be abnormally short axonemal projections and (at 
least for osm-1) an assembly of ectopic doublet microtubules which 
constitute cilia-like posterior projections.  These animals have 
various behavioral abnormalities which presumably result from a 
failure of the sensory cilia to project into the environment (i.e.  
daf, osm, che etc.).
To understand how these genes function, we have begun a molecular 
characterization of the two loci.  We obtained from Don Riddle's lab 
alleles osm-1(m530), 8), and daf-10(m534), which 
were generated in RW7097.  Spontaneous revertants were identified and, 
they and the mutants from which they were derived were outcrossed.  
DNA samples from outcrossed mutants, revertants, and recombinants were 
isolated for Southern analysis.  For each gene, a single Tc1-
containing EcoRI fragment which co-segregated with the mutant 
phenotype was identified.  These EcoRI fragments were cloned and their 
identities verified.  DNA fragments flanking the Tc1 elements were 
used to screen a genomic library provided by Chris Link.  From these 
experiments we identified three overlapping lambda clones for osm-1 
and a single lambda clone which hybridized to the daf-10 probe.  The 
osm-1 and daf-10 lambda clones were sent to John Sulston and Alan 
Coulson at the MRC for assignment on the physical map.  Both genes 
landed in large contigs.  The daf-10 clone was placed within 
approximately 20kb and to the right of fem-3, and the osm-1 clone was 
placed within about 100kb and to the right of mtl-1.  In addition 
fragments of the genomic clones were used to identify hybridizing cDNA 
clones from a library provided by Stuart Kim.  The osm-1 and daf-10 
probes each identified cDNAs.  We have begun sequencing these clones 
and using them as probes for northern analyses.