--Bruce Veit, Erik Vollbrecht and Sarah Hake

Kn1-O and Kn1-2F11 are two dominant mutations, both closely linked to Adh1, that interfere with the differentiation of vascular cells in the leaf blade. Recently, DNA associated with the knotted allele Kn1-2F11 was cloned by its association with the previously cloned transposable element, Ds2. Expression of the knotted phenotype in Kn1-2F11 depends on the presence of Ds2; excision of Ds2 results in reversion to a non-knotted phenotype (Hake, Vollbrecht and Freeling, in press).

We have used the Kn1-2F11 clone as a hybridization probe to examine the organization of homologous DNA sequences associated with Kn1-O. In comparisons of knotted vs. normal siblings, no insertion sequences comparable to the Ds2 of Kn1-2F11 were found; however, by Southern analysis, this region appears to be duplicated in Kn1-O. Cloning of junction fragments unique to Kn1-O indicates the tandem duplication of a 19kb region. Kn1-Ox, an extremely knotted derivative of Kn1-O, shows a triplication of this same region, while Kn1-Od, a normal derivative of Kn1-O, is missing the region.

Studies are in progress to determine if novel transcripts associated with the unique duplication junction might be responsible for the knotted phenotype. The importance of this region is suggested by at least two separate instances in which the insertion of Mu related transposable elements within 1kb of the duplication junction is associated with the loss of the knotted phenotype. The junction is also implicated by its proximity (<2kb) to the site at which the insertion of Ds2 in Kn1-2F11 produces a knotted phenotype.

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