Worm Breeder's Gazette 10(2): 60

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Identification of a zyg-11 Clone

Philip W. Carter and Kenneth J. Kemphues

Figure 1

The zyg-11 gene, on chromosome II, is required for proper 
cytoplasmic organization in early embryos.  Mutations in the gene have 
a strict maternal-effect lethal phenotype.  In embryos from zyg-11 
mothers, meiosis arrests at metaphase II, there is a delay in the 
appearance of pronuclei, the position of the first cleavage furrow is 
abnormal, and P granules fail to segregate properly (Kemphues et al., 
Dev.  Biol.  113, 449 (1986)).  We believe that a molecular analysis 
of this gene will help us to understand the nature and function of 
maternally-encoded factors in the egg.  To identify a zyg-11 clone, we 
made use of a 400kb contig from Alan Coulson and John Sulston that we 
thought likely to contain zyg-11.  We had previously shown that the 
left-most cosmid of the contig overlapped the left end of mnDf103, and 
the remainder of the contig extended to the right on the genetic map (
toward zyg-11).  The contig also covered the sites of 2 Tc1 elements 
identified in a mutator strain by Ikue Mori, Don Moerman and Bob 
Waterston (WBG 10 #1, p.47).  Ikue kindly sent us a worm strain 
containing the two Tc1 's, and we showed by 3-factor crosses that zyg-
11 mapped to the left of Tc1 #40 (RW#200L).  Therefore, zyg-11 had to 
be in the contig somewhere.  We decided to find the position of zyg-11 
in the contig by using individual cosmids as probes of a Southern blot 
containing DNA from 2 mut-2 induced zyg-11 mutants (it50 and it65) 
that we had found earlier.  The first cosmid that we tested, DH11, 
must contain at least part of zyg-11 since it detected DNA 
rearrangements in both it50 and it65.  The rearrangement in it50 
appeared to be a 2.9kb insertion, while the rearrangement in it65 
appeared to be a 1.6kb insertion.  Both insertions occurred within a 3.
0kb EcoRI fragment.  Additional Southern analysis narrowed down the 
points of insertion to an 800-base EcoRI-BglII fragment.  We are now 
sequencing that fragment.  We will also use the clone as a probe to 
study transcription of zyg-11.  The cosmid DH11 also seems to cover an 
endpoint of mnDf104 since it detects a novel restriction fragment when 
used as a probe against mnDf104 DNA.  The figure below shows some 
clones of the contig, along with some genetic markers.
[See Figure 1]

Figure 1