Location: Vegetable Crops ResearchTitle: Mining and validation of pyrosequenced simple sequence repeats (SSRs) from American cranberry (Vaccinium macrocarpon Ait.) Author
Submitted to: Theoretical and Applied Genetics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/17/2011
Publication Date: 1/2/2012
Citation: Zalapa, J.E., Simon, P.W., Hummer, K.E., Bassil, N.V., Senalik, D.A., Zhu, H., Mccown, B.H., Zeldin, E., Speers, J., Hyman, J. 2012. Mining and validation of pyrosequenced simple sequence repeats (SSRs) from American cranberry (Vaccinium macrocarpon Ait.). Theoretical and Applied Genetics. 124(1):87-96. Interpretive Summary: Cranberries are a major commercial fruit crop in North America, but limited genetic resources have been developed for the species. Furthermore, the paucity of genetic tools has hampered the advance of genetic research in cranberry and other closely related species. Therefore, we used new and powerful DNA sequencing technologies to produce hundreds of thousands of cranberry ‘HyRed’ sequences were we discovered a total 107,244 genetic markers that could potentially be used in cranberry genetic research. Cranberry cultivars are clonally preserved and propagated with established beds remaining productive for over 100 years. Thus, the development of efficient, reliable methods for the variety identification will be vital to the success of the cranberry industry. Here we report the developed 48 cranberry markers that were very useful to differentiate a sample including widely used cranberry cultivars and for parentage identification in breeding cranberry breeding programs. New sequencing technologies were a cost-effective and efficient away to identify numerous cranberry markers that will likely lead to innovative strategies to speed up the breeding of unique cranberry cultivars to meet the current and future challenges of an important crop for American industry.
Technical Abstract: The American cranberry (Vaccinium macrocarpon Ait.) is a major commercial fruit crop in North America, but limited genetic resources have been developed for the species. Furthermore, the paucity of DNA markers has hampered the advance of genetic research in cranberry and the Ericaceae family in general. Therefore, we used Roche 454 sequencing technology to perform genome-wide shotgun sequencing of the cranberry cultivar ‘HyRed’. After de novo assembly, the obtained sequence covered 266.3Mb of the estimated 540 - 590 Mb in cranberry genome. A total of 107,244 SSR loci were detected with an overall density across the genome of 403 SSR/Mb. The AG repeat was the most frequent motif in cranberry accounting for 35% of all SSRs and together with AAG and AAAT accounted for 46% of all loci discovered. In order to validate the SSR loci, we designed 96 primer-pairs using contig sequence data containing perfect SSR repeats, and studied the genetic diversity of among 25 cranberry genotypes. We indentified 48 polymorphic SSR loci with 2 to 15 alleles per locus for a total of 323 alleles in the 25 cranberry genotypes. Genetic clustering by principal coordinates and genetic structure analyses confirmed the heterogeneous nature of cranberries. The parentage composition of several hybrid cultivars was clearly evident from our structure analyses. Whole genome shotgun 454 sequencing was a cost-effective and efficient away to identify numerous SSR repeats in the cranberry sequence for marker development.