Originally published as Genetics Published Articles Ahead of Print on December 6, 2006.

Genetics, Vol. 175, 981-992, March 2007, Copyright © 2007
doi:10.1534/genetics.106.066837

Generating Novel Allelic Variation Through Activator Insertional Mutagenesis in Maize

Ling Bai^*, Manjit Singh^,1, Lauren Pitt, Meredith Sweeney and Thomas P. Brutnell^*,,2

^* Department of Plant Biology, Cornell University, Ithaca, New York 14853 and Boyce Thompson Institute, Cornell University, Ithaca, New York 14853

² Corresponding author: Boyce Thompson Institute, Cornell University, 1 Tower Rd., Ithaca, NY 14853.
E-mail: tpb8{at}cornell.edu

The maize transposable element Activator (Ac) has been exploited as an insertional mutagen to disrupt, clone, and characterize genes in a number of plant species. To develop an Ac-based mutagenesis platform for maize, a large-scale mutagenesis was conducted targeting the pink scutellum1 locus. We selected 1092 Ac transposition events from a closely linked donor Ac, resulting in the recovery of 17 novel ps1 alleles. Multiple phenotypic classes were identified corresponding to Ac insertions in the 5'-UTR and coding region of the predicted Ps1 gene. To generate a stable allelic series, we employed genetic screens and identified 83 germinally heritable ps1 excision alleles. Molecular characterization of these excision alleles revealed a position-dependent bias in excision allele frequencies and the predominance of 7- and 8-bp footprint products. In total, 19 unique ps1 excision alleles were generated in this study, including several that resulted in weak mutant phenotypes. The analysis of footprint alleles suggests a model of Ac excision in maize that is consistent with recent in vitro studies of hAT element excision. Importantly, the genetic and molecular methods developed in this study can be extended to generate novel allelic variation at any Ac-tagged gene in the genome.

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