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ARS Home » Plains Area » Fargo, North Dakota » Edward T. Schafer Agricultural Research Center » Sunflower and Plant Biology Research » Research » Publications at this Location » Publication #146240
Title: ASSESSING GENETIC VARIABILITY AMONG SIXTEEN PERENNIAL HELIANTHUS SPECIES USING PCR-BASED TRAP MARKERS

Author
item Hu, Jinguo
item Seiler, Gerald
item Jan, Chao-Chien
item Vick, Brady

Submitted to: Proceedings Sunflower Research Workshop
Publication Type: Proceedings
Publication Acceptance Date: 2/12/2003
Publication Date: 3/1/2003
Citation: HU, J., SEILER, G.J., JAN, C.C., VICK, B.A. ASSESSING GENETIC VARIABILITY AMONG SIXTEEN PERENNIAL HELIANTHUS SPECIES USING PCR-BASED TRAP MARKERS. PROCEEDINGS SUNFLOWER RESEARCH WORKSHOP. 2003. www.sunflowernsa.com/research_statistics/research_workshop/documents/88.pdf

Interpretive Summary: Wild species are important reservoirs of useful genes for crop improvement. The wild species of sunflower are a valuable genetic resource for improving the cultivated sunflower crop. The majority of wild sunflower species are perennial, presenting a special challenge if they are to be used in a genetic enhancement program for the annual crop species. However, the perennial species have been particularly useful as a source of resistance genes for several of the major sunflower pathogens, especially downy mildew. The identification of genes from wild species can be very challenging because of the genetic diversity and complexity within each species' genome. An effective and efficient molecular marker technique would help facilitate the transfer of desirable agronomic genes from the wild ancestors to the sunflower crop. In our laboratory, we developed a targeted region amplified polymorphism (TRAP) marker system to generate DNA markers. This system uses a fixed primer complementary to the known gene sequences in the sunflower expressed sequence tag (EST) database and a second primer with a random sequence. The objective of this study was to present preliminary results assessing the genetic variability of the markers among 16 Helianthus species and two perennial hybrids using TRAP markers. Since we are interested in the disease resistance genes in the wild species, we designed the fixed primer to target a sequence coding the leucine rich repeat (LRR)-like protein in the sunflower EST database. It is well documented that the LLR -like protein is an essential component of the plants defense system to diseases. Our preliminary results suggest that TRAP markers can be used to assess the genetic diversity of the wild perennial sunflower species. The perennial species studied were highly polymorphic at the DNA level as revealed by the three TRAP primers, one of which is targeting a gene sequence involved in disease resistance. Thus, these species are particularly beneficial to a breeding project searching for resistance to new races of pathogens. However, the association between the variation at the DNA level and the reaction to a particular pathogen will need to be investigated further.

Technical Abstract: The wild species of the genus Helianthus are a valuable genetic resource for improving cultivated sunflower. They have been particularly useful as a source of resistance genes for several of the major sunflower pathogens, especially downy mildew caused by Plasmopara halstedii. The identification of genes from wild species can be very challenging because of the genetic diversity and complexity within each species' genome. An effective and efficient molecular marker technique would help facilitate the transfer of desirable agronomic genes from the wild ancestors to the sunflower crop. The TRAP (target region amplified polymorphism) marker system developed in our laboratory was used to generate DNA markers. This system, modified from the SRAP (sequence-related amplified polymorphism) marker system, uses a fixed primer complementary to the known gene sequences in the sunflower EST (expressed sequence tag) database and a second primer of random sequence in the PCR reaction. The objective of this study was to present preliminary results assessing the genetic variability of PCR-based markers among 16 Helianthus species and two perennial hybrids using PCR-based TRAP markers. Since we are interested in the disease resistance genes in the wild species, we designed the fixed primer to target a sequence coding the LRR (leucine rich repeat)-like protein in the sunflower EST database. Forty out of the 42 DNA samples produced very good amplification. The fragments sized from 50 to 900 base pairs, separated well on a 6.5% polyacrylamide sequencing gel. The NTSYSpc software clustered the species together and generated a phylogenetic tree. The cultivated annual lines clustered into a distinct group. The perennial species of the Ciliares section were separated into a separate cluster, following the classical taxonomic classification for this section. The TRAP markers grouped the five populations of H. mollis into the same cluster, indicating that the markers are amplified in the individuals and in different populations of the same species. In the present study, many of the perennial species belong to the Divaricati section, which contains many species. In general, the TRAP markers grouped the species into somewhat similar clusters, much the same as the classical taxonomic system. Our preliminary results suggest that TRAP markers can be used to assess the genetic diversity of the wild perennial sunflower species. The perennial species studied were highly polymorphic at the DNA level as revealed by the three TRAP primers, one of which is targeting a gene sequence involved in disease resistance. Thus, these species are particularly beneficial to a breeding project searching for resistance to new races of pathogens. However, the association between the variation at the DNA level and the reaction to a particular pathogen needs to be investigated further.