|LIU, PINGWU - COLLABORATOR|
|RICHARD JR, EDWARD|
Submitted to: American Society of Sugar Cane Technologists
Publication Type: Abstract Only
Publication Acceptance Date: 6/2/2010
Publication Date: 6/16/2010
Citation: Liu, P., Pan, Y.-B., Grisham, M.P., Tew, T.L., White, W.H., Richard Jr, E.P. 2010. Searching for DNA markers associated with sugar content in sugarcane [abstract]. Journal of the American Society of Sugar Cane Technologists. 30:138.
Technical Abstract: Sugar content is among the most important traits in sugarcane for its commercial production. A relatively stable genetic character in sugarcane breeding programs, sugar content requires direct instrumental measurements during selection. This process is labor intensive and must be done only when the crop reaches maturity. The objective of this study was to explore the possibility of identifying suitable DNA markers for marker-assisted selection of sugar content in sugarcane. We used the candidate gene approach for marker development. From the GenBank database, we fetched four genomic sequences encoding for SuSy (sucrose synthase) and SPS (sucrose phosphate synthase) and 76 expressed sequence tags (ESTs) including seven for SPS, 41 for SuSy, and 28 for AInv (acid invertase). These three enzymes play important roles in sugar metabolism in sugarcane. We also found homologous sequences of these three genes of Sorghum, Oryza sativa, and Arabidopsis. Sequences belonging to the same gene were aligned using DNAMAN® software to produce homology trees. Based on these homolog trees, 14 ESTs or whole genomic sequences were chosen for the design of 16 pairs of PCR primers. The primer pairs were used to amplify PCR products from the genomic DNA of six clones from a mapping population derived from selfing pollination of LCP 85-384. These clones were segregating for sugar content but were similar for other phenotypic traits. The amplified PCR products were separated by electrophoresis in 1.5% agarose gels, stained with ethidium bromide, visualized under UV lights, and photographically documented. Each of 14 primer pairs produced a single DNA band as expected and two primer pairs did not produce any band. The DNA products of crude PCR reaction mixtures were cloned directly into a pCR4-TOPO vector from Invitrogen. Four to six recombinant clones from each PCR reaction were sequenced. Analyses of the DNA sequence data demonstrated the following: 1) the amplified PCR products from any sample were mixtures of DNA fragments displaying sequence variability, very likely due to the aneu-polyploidy nature of sugarcane; and 2) the sequence variability of LCP 85-384 was transmitted to its progeny. We will further scrutinize the variability of these DNA sequences to determine if there is any variability associated with sugar content.