Utilizing single primers as molecular markers in Poa spp

Authors: Kindiger, Bryan; CONLEY, TERRY - OKLA. CITY UNIV.

Submitted to: Journal of Japanese Society of Grassland Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/9/2009
Publication Date: 10/1/2009
Citation: Kindiger, B.K., Conley, T. 2009. Utilizing single primers as molecular markers in Poa spp. Japanese Society of Grassland Science. 55:206-215.

Interpretive Summary: The development of DNA marker systems for grass forages is restrictive due to the high cost and the time-consuming nature of marker development. However, if molecular markers were available, they could have direct and immediate application in selection and breeding studies. The development of a rapid and inexpensive molecular marker approach that builds upon prior marker development in other grass species could reduce the cost and the time consuming nature of marker development. In addition, an approach that would take advantage of duplicated and inverted DNA regions that comprise the genome composition of many grass forages, could generate molecular markers that exhibit a high degree of specificity. This research presents a simple and effective approach, utilizing presently available molecular markers from other grass forage and cereal grass species, to identify inverted repeat DNA sequence regions in bluegrass. The application and value of the generated markers were further examined and evaluated across eight bluegrass species and three of their hybrids. The study indicates that the duplicated and inverted DNA sequence regions in bluegrass, are highly specific to individuals and across species and this attribute allows these markers to be useful for DNA fingerprinting and cultivar identification. The results of this study will be of value to researchers working with species possessing complex genomes.

Technical Abstract: We describe an approach, utilizing single SSR primers at high annealing temperature, that identifies palindrome or quasi-palindrome regions in polyploid genomes. Palindromes or quasi-palindromes are unique genome sequences considered to be potentially abundant in polyploid plant genomes and the development of a marker approach that could identify such sequences, could have desirable applications in studies utilzing polyploid species. Similar to RAPD-PCR or AP-PCR, this approach requires no prior genome information, utilizes agarose gels and can be visualized with ethidium bromide. The approach was evaluated in Poa arachnifera and various Poa interspecific hybrids and was determined to be effective in identifying markers that coexpress in the examined materials. DNA sequence analysis of the PCR amplification products also identified a wide array of potentially homeologous loci within and among the related individuals. The study resulted in the identification of 157 single primers that are useful in identifying informative polymorphisms across P. arachnifera and its interspecific hybrids. Preliminary evaluations of additional Poa spp., Bromus inermis, Dactylis glomerata, Thinopyrum ponticum and Aragostis spp. suggest wide utility of this approach toward genotyping additional polyploid grass species. The results of this study will be of value to researchers working with species possessing complex genomes.