Molecular genetics of the Caenorhabditis elegans heterochronic gene lin-14.
We describe a general strategy for the genetic mapping in parallel of multiple restriction fragment length polymorphism (RFLP) loci. This approach allows the systematic identification for cloning of physical genetic loci within about 100 kb of any gene in Caenorhabditis elegans. We have used this strategy of parallel RFLP mapping to clone the heterochronic gene lin-14, which controls the timing and sequence of many C. elegans postembryonic developmental events. We found that of about 400 polymorphic loci in the C. elegans genome associated with the Tc1 family of repetitive elements, six are within 0.3 map unit of lin-14. The three closest lin-14-linked Tc1-containing restriction fragments were cloned and used to identify by hybridization an 830-kb region of contiguous cloned DNA fragments assembled from cosmid and yeast artificial chromosome libraries. A lin-14 intragenic recombinant that separated a previously cryptic lin-14 semidominant mutation from a cis-acting lin-14 suppressor mutation was used to map the location of the lin-14 gene to a 25-kb region of this 830-kb contig. DNA probes from this region detected lin-14 allele-specific DNA alterations and a lin-14 mRNA. Two lin-14 semi-dominant alleles, which cause temporally inappropriate lin-14 gene activity and lead to the reiterated expression of specific early developmental events, were shown to delete sequences from the lin-14 gene and mRNA. These deletions may define cis-acting sequences responsible for the temporal regulation of lin-14.
Ruvkun
G
G
Department of Molecular Biology, Massachusetts General Hospital, Boston.
Ambros
V
V
Coulson
A
A
Waterston
R
R
Sulston
J
J
Horvitz
H R
HR
D000483:Alleles; D000818:Animals; D002107:Caenorhabditis / Q000235:genetics*; D003001:Cloning, Molecular; D005796:Genes; D009154:Mutation; D012150:Polymorphism, Restriction Fragment Length; D012333:RNA, Messenger / Q000235:genetics; D011995:Recombination, Genetic; D015183:Restriction Mapping; D014158:Transcription, Genetic
D012333:RNA, Messenger