Worm Breeder's Gazette 15(4): 39 (October 1, 1998)

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.

Sequence analysis of lin-18 mutants

Wendy S. Katz, Helieh S. Oz, Jeff Goodwin

Department of Biochemistry, University of Kentucky College of Medicine. Lexington KY 40536-0298

Our laboratory is investigating the role of lin18 in vulval cell
orientation.  Worms mutant for lin-18 generate P7.p lineages that are
misoriented, forming a pseudovulva posterior to a defective vulva (WBG

We cloned a gene that rescues lin-18 mutants in germline transformation
experiments (WBG 15(1):60).  The rescuing gene encodes a predicted
receptor tyrosine kinase.  To confirm that this gene is lin-18, we
amplified the gene from lin-18 mutants and  sequenced it.  We sequenced
products of at least two independent PCR reactions for each allele. 
Regions that contained a sequence change were sequenced on top and
bottom strands for each allele.

The reference allele, e620, contained a C to T change.  This converts a
Gln codon to a stop codon at position 84 in the amino acid sequence
predicted from cDNA analysis (see below).  ga75 contained a 380 bp
internal deletion in the predicted extracellular domain.  The deletion
removes the fourth exon (codons 85-136) and causes a shift in reading
frame.  This is predicted to generate a truncated gene product ending in
a novel sequence of 13 amino acids following codon 84.

To confirm the predicted splice pattern reported in ACeDB, we are
cloning and sequencing cDNAs.  In the first clone analyzed, we found
that the splice pattern was not as predicted.  The cDNA sequence we
obtained is consistent with the e620 sequence analysis, as it places the
mutation in an exon; the original splice prediction resulted in the e620
point mutation being in an intron.  We are currently examining
independent cDNA clones to confirm this result.