GENETIC IMPROVEMENT OF SUGARCANE BY CONVENTIONAL AND MOLECULAR APPROACHES
Location: Sugarcane Research Unit
Title: Identification of RAPD marker associated with brown rust resistance in sugarcane
Submitted to: American Society of Sugar Cane Technologists
Publication Type: Abstract Only
Publication Acceptance Date: June 2, 2010
Publication Date: June 16, 2010
Citation: Muhammad, K., Pan, Y.-B., Grisham, M.P., Iqbal, J. 2010. Identification of RAPD marker associated with brown rust resistance in sugarcane [abstract]. Journal of the American Society of Sugar Cane Technologists. 30:138.
Susceptibility to brown rust caused by Puccinia melanocephala is a major reason for the withdrawal of sugarcane cultivars from production. An efficient way to control the disease is to breed cultivars with durable resistance. Our aim was to identify random amplified polymorphic DNA (RAPD) markers that associate closely with brown rust resistance using a mapping population derived from self-pollination of the Louisiana cultivar LCP 85-384. Two bulked DNA samples were generated, one from five clones that were rated highly resistant (R) and the other from five clones rated highly susceptible (S) to the brown rust based on field ratings of disease symptoms. Additionally, the 10 clones were selected because they shared several other phenotypic traits, i.e., plant height, stalk number, sugar and fiber content, etc. A total of 319 RAPD primers were screened by polymerase chain reaction (RAPD-PCR) on the bulked DNA samples along with the DNA of LCP 85-384 (positive control) and water (negative) samples. The amplified RAPD-PCR products were separated by electrophoresis in 1.5% agarose gels, stained with ethidium bromide, visualized under UV lights, and photographically documented. Although 312 primers produced similar banding patterns, seven primers, i.e., AM-14, AM-19, L-18, N-17, Y-5, AO-15, and N-6, produced DNA bands that were discriminative between the resistant and susceptible bulked samples. RAPD-PCR reactions were repeated three times on these seven primers to validate these findings. One of these discriminative RAPD-PCR products, a 450 base pair-long DNA band, from the primer N-06 was produced from all five resistant clones as well as LCP 85-384, but not from the five susceptible clones. We are in the process of cloning and sequencing this DNA band to begin designing a sequenced characterized amplified region (SCAR) marker that is reliable and reproducible.