Worm Breeder's Gazette 14(2): 27 (February 1, 1996)

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.

Daring rescue! Corruption! in.... CLONEWATCH

Tom Barnes, Julia Pak, Athena Vouloumanos, Siegfried Hekimi

Department of Biology, McGill University, Montreal H3A1B1, PQ, Canada

In the course of narrowing the unc-14 rescuing region to a 5.2 kb Hind
III - Sal I fragment that is mostly occupied by part of the YK363 gene
(see also Ogura and Ohshima WBG 14(1) p51), we came across two unusual
clones.  We had established a restriction map across a 60 kb region,
with particular attention to Eco RI sites, and had obtained 8 clones
from this region from the LMB, which we mapped and tested for rescue. 
The first unusual clone, P1a74 (which uses a P1 phage-derived vector),
was a clone which should have rescued.  This clone has a wild-type
restriction map, and the 11 kb Eco RI fragment which contains unc-14(+)
is in the middle of this clone.  However, 3/3 lines (and 18 F1 Rol)
carrying this clone were not rescued, not even partially (since Unc-14
adults are strongly paralyzed this is a sensitive test).  This suggests
that the clone is not faithful to the genomic sequence and contains a
mutation which completely inactivates unc-14(+).  Of course this could
just be a sporadic case, but we wonder if anyone else has noticed any
odd behaviour of P1 clones?

The other clone was cosmid K06F6.  This clone rescues, but appeared on
first inspection to be a chimera.  It is a full-length cosmid and
contains 7 contiguous Eco RI fragments from the unc-14 region spanning
24.5 kb, but at least 4 fragments clearly do not come from this region. 
There are also at least 4 Hind III fragments in this clone not shared by
overlapping clones.  The original fingerprint of this clone, however,
does not appear problematic (Alan Coulson, pc).  The simplest
explanation is that this clone has rearranged without an apparent net
loss of DNA.  Bizarre.