Molecular cloning of lin-29, a heterochronic gene required for the differentiation of hypodermal cells and the cessation of molting in C.elegans

The lin-29</it> gene product of C.elegans activates a temporal developmental switch for hypodermal cells. Loss-of-functlon lin-29</it> mutations result in worms that fail to execute a stage-specific pattern of hypodermal differentiation that includes exist from the cell cycle, repression of larval cuticle genes, activation of adult cuticle genes, and the cessation of molting. Combined genetic and physical mapping of restriction fragment length polymorphisms (RFLPs) was used to identify the lin-29</it> locus. A probe from the insertion site of a Tc1 (maP1), closely linked and to the left of lin-29</it> on the genetic map, was used to identify a large set of overlapping cosmid, lambda and yeast artificial chromosome (YAC) clones assembled as part of the C.elegans physical mapping project. Radiolabeled DNA from one YAC clone identified two distinct allele-specific alterations that cosegregated with the lin-29</it> mutant phenotype in lin-29</it> intragenic recombinants. lin-29</it> sequences were severely under-represented in all cosmid and lambda libraries tested, but were readily cloned in a YAC vector, suggesting that the lin-29</it> region contains sequences incompatible with standard prokaryotic cloning techniques.