Characterisation of single nucleotide polymorphisms in sugarcane ESTs
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Commercial sugarcane cultivars (Saccharum spp. hybrids) are both polyploid and aneuploid with chromosome numbers in excess of 100; these chromosomes can be assigned to 8 homology groups. To determine the utility of single nucleotide polymorphisms (SNPs) as a means of improving our understanding of the complex sugarcane genome, we developed markers to a suite of SNPs identified in a list of sugarcane ESTs. Analysis of 69 EST contigs showed a median of 9 SNPs per EST and an average of 1 SNP per 50 bp of coding sequence. The quantitative presence of each base at 58 SNP loci within 19 contiguous sequence sets was accurately and reliably determined for 9 sugarcane genotypes, including both commercial cultivars and ancestral species, through the use of quantitative light emission technology in pyrophosphate sequencing. Across the 9 genotypes tested, 47 SNP loci were polymorphic and 11 monomorphic. Base frequency at individual SNP loci was found to vary approximately twofold between Australian sugarcane cultivars and more widely between cultivars and wild species. Base quantity was shown to segregate as expected in the IJ76-514 × Q165 sugarcane mapping population, indicating that SNPs that occur on one or two sugarcane chromosomes have the potential to be mapped. The use of SNP base frequencies from five of the developed markers was able to clearly distinguish all genotypes in the population. The use of SNP base frequencies from a further six markers within an EST contig was able to help establish the likely copy number of the locus in two genotypes tested. This is the first instance of a technology that has been able to provide an insight into the copy number of a specific gene locus in hybrid sugarcane. The identification of specific and numerous haplotypes/alleles present in a genotype by pyrophosphate sequencing or alternative techniques ultimately will provide the basis for identifying associations between specific alleles and phenotype and between allele dosage and phenotype in sugarcane.
This research was undertaken with funding from the Cooperative Research Centre for Sugar Industry Innovation through Biotechnology.
- Aitken K, Jackson P, Piperidis G, McIntyre L (2004) QTL identified for yield components in a cross between a sugarcane (Saccharum spp.) cultivar Q165A and a S. officinarum clone IJ76-514. In: Proceedings for the 4th international crop science congress, Brisbane, Australia, 26 September–1 October 2004. http://www.cropscience.org.au
- Bhattramakki D, Rafalski A (2001) Discovery and application of single nucleotide polymorphism markers in plants. In: Henry RJ (ed) Plant genotyping: the DNA fingerprinting of plants. CAB International, Lismore, pp 179–191Google Scholar
- Casu RE, Dimmock CM, Chapman SC, Grof CPL, McIntyre CL, Bonnett GD, Manners JM (2004) Identification of differentially expressed transcripts from maturing stem of sugarcane by in silico analysis of stem expressed sequence tags and gene expression profiling. Plant Mol Biol 54:503–517CrossRefPubMedGoogle Scholar
- McIntyre CL, Jackson M, Cordeiro G, Amouyal O, Eliott F, Henry RJ, RE Casu, Hermann S, Aitken KS, Bonnett GD (2006) The identification and characterisation of alleles of sucrose phosphate synthase gene family III in sugarcane. Mol Breed (in press)Google Scholar
- Mototani H, Mabuchi A, Saito S, Fujioka M, Iida A, Takatori Y, Kotani A, Kubo T, Nakamura K, Sekine A, Murakami Y, Tsunoda T, Notoya K, Nakamura Y, Ikegawa S (2005) A functional single nucleotide polymorphism in the core promoter region of CALM1 is associated with hip osteoarthritis in Japanese. Hum Mol Genet 14:1009–1017CrossRefPubMedGoogle Scholar
- Pfost DR, Boyce-Jacino MT, Grant DM (2000) A SNPshot: pharmacogenetics and the future of drug therapy. TIBTECH 18:334–338Google Scholar
- Ronaghi M (2001) Pyrosequencing sheds light on DNA sequencing. J Chromatogr B 739:345–355Google Scholar
- Rossi M, Araujo PG, Paulet F, Garsmeur O, Dias VM, Chen H, Van Sluys M-A, D’Hont A (2003) Genomic distribution and characterization of EST-derived resistance gene analogs (RGAs) in sugarcane. Mol Gen Genet 269:406–419Google Scholar
- Tishkoff SA, Goldman A, Calafell F, Speed WC, Deinard AS, Bonne-Tamir B, Kidd JR, Pafstis AJ, Jenkins T, Kidd KK (1998) A global haplotype analysis of the myotonic dystrophy locus: implications for the evolution of modern humans and for the origin of myotonic dystrophy mutations. Am J Hum Genet 62:1389–1402CrossRefPubMedGoogle Scholar
- Waters DLE, Henry RJ, Reinke RF, Fitzgerald MA (2005) Gelatinisation temperature of rice explained by polymorphisms in starch synthase. Plant Biotechnol J (In press)Google Scholar