High-throughput linkage analysis of Mutator insertion sites in maize

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Yi, Gibum
Luth, Diane
Goodman, Timothy
Lawrence, Carolyn
Becraft, Philip
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The Department of Agronomy seeks to teach the study of the farm-field, its crops, and its science and management. It originally consisted of three sub-departments to do this: Soils, Farm-Crops, and Agricultural Engineering (which became its own department in 1907). Today, the department teaches crop sciences and breeding, soil sciences, meteorology, agroecology, and biotechnology.

The Department of Agronomy was formed in 1902. From 1917 to 1935 it was known as the Department of Farm Crops and Soils.

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  • Department of Farm Crops and Soils (1917–1935)

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Genetics, Development and Cell Biology

The Department of Genetics, Development, and Cell Biology seeks to teach subcellular and cellular processes, genome dynamics, cell structure and function, and molecular mechanisms of development, in so doing offering a Major in Biology and a Major in Genetics.

The Department of Genetics, Development, and Cell Biology was founded in 2005.

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Bioinformatics and Computational Biology
The Bioinformatics and Computational Biology (BCB) Program at Iowa State University is an interdepartmental graduate major offering outstanding opportunities for graduate study toward the Ph.D. degree in Bioinformatics and Computational Biology. The BCB program involves more than 80 nationally and internationally known faculty—biologists, computer scientists, mathematicians, statisticians, and physicists—who participate in a wide range of collaborative projects.
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Insertional mutagenesis is a cornerstone of functional genomics. High-copy transposable element systems such as Mutator (Mu) in maize (Zea mays) afford the advantage of high forward mutation rates but pose a challenge for identifying the particular element responsible for a given mutation. Several large mutant collections have been generated in Mu-active genetic stocks, but current methods limit the ability to rapidly identify the causal Mu insertions. Here we present a method to rapidly assay Mu insertions that are genetically linked to a mutation of interest. The method combines elements of MuTAIL (thermal asymmetrically interlaced) and amplification of insertion mutagenized sites (AIMS) protocols and is applicable to the analysis of single mutants or to high-throughput analyses of mutant collections. Briefly, genomic DNA is digested with a restriction enzyme and adapters are ligated. Polymerase chain reaction is performed with TAIL cycling parameters, using a fluorescently labeled Mu primer, which results in the preferential amplification and labeling of Mu-containing genomic fragments. Products from a segregating line are analyzed on a capillary sequencer. To recover a fragment of interest, PCR products are cloned and sequenced. Sequences with lengths matching the size of a band that co-segregates with the mutant phenotype represent candidate linked insertion sites, which are then confirmed by PCR. We demonstrate the utility of the method by identifying Mu insertion sites linked to seed-lethal mutations with a preliminary success rate of nearly 50%.


This article is from The Plant Journal 58 (2009): 883–892, doi:10.1111/j.1365-313X.2009.03821.x.