Allelic divergence and cultivar-specific SSR alleles revealed by capillary electrophoresis using fluorescence-labeled SSR markers in sugarcane

Authors: CHANDRA, AMARESH - Collaborator; Grisham, Michael; Pan, Yong-Bao

Submitted to: Genome
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/17/2014
Publication Date: 8/26/2014
Publication URL: http://handle.nal.usda.gov/10113/59717
Citation: Chandra, A., Grisham, M.P., Pan, Y.-B. 2014. Allelic divergence and cultivar-specific SSR alleles revealed by capillary electrophoresis using fluorescence-labeled SSR markers in sugarcane. Genome. 57:363–372. doi:10.1139/gen-2014-0072.

Interpretive Summary: India maintains the largest collection of sugarcane genetic resources in the world. To maximize its utilization in sugarcane improvement, this collection needs to be genetically characterized both phenotypically and molecularly. However, due to the co-existence of multiple sets of chromosomes, or polyploidy, in cells of sugarcane plant, precise detection and size determination of multiple DNA fragments using conventional gel electrophoresis methods have been a technical challenge. This study used a capillary electrophoresis-based detection system in conjunction with fluorescence-labeling of DNA fragments to fingerprint 24 sugarcane cultivars from India and the U.S. In total, 213 DNA fragments were detected and the molecular sizes of these DNA fragments determined precisely upon calibration against 16 DNA size standards. One hundred sixty-one DNA fragments were common to both Indian and the U.S. sugarcane cultivars, 25 DNA fragments were found only in the Indian cultivars, and 27 DNA fragments were found only in the U.S. cultivars. Based on the genetic divergence among these DNA fragments, genetic relatedness computer software DNAMAN placed the 24 sugarcane cultivars into three distinctive groups. Groups I and II were made up of Indian cultivars and Group III of the U.S. cultivars. Of particular interest was the identification of six DNA fragments that were in the Indian cultivars only. The results from this study provide molecular genetic information for Indian sugarcane breeders in areas of cultivar identification, cross fidelity assessment, and germplasm utilization.

Technical Abstract: Though sugarcane cultivars (Saccharum spp. hybrids) are complex aneu-polyploid hybrids, genetic evaluation and tracking of clone- or cultivar-specific alleles become possible due to capillary electrophoregrams (CE) using fluorescence-labeled SSR primer pairs. Twenty-four sugarcane cultivars, 12 each from India and the U.S., were genetically assessed using 21 fluorescence-labeled polymorphic microsatellite markers. These markers primed the amplification of 213 alleles. Of these alleles, 161 were common to both Indian and the U.S. cultivars, 25 were specific to the Indian cultivars, and 27 were observed only in the U.S. cultivars. Only 10 alleles were monomorphic among all 24 cultivars. A high level of heterozygosity was observed in both Indian (159 alleles, or 82.4%) and the U.S. (154 alleles, or 91.1%) cultivars resulting in an average PIC values of 0.66 and 0.77 and marker index (MI) values of 5.07 and 5.58, respectively. The Pearson correlation between PIC and MI was found to be significant in both sets of cultivars (r = 0.58 and 0.69). The UPGMA clustering dendrogram separated the Indian and the U.S. cultivars into three distinct clusters at 59% homology level. These results propose the potential utility of six Indian cultivar-specific SSR alleles [mSSCIR3_182 (specific to CoJ64), SMC486CG_229, SMC36BUQ_125, and mSSCIR74_216 (specific to CoLk8001), SMC334BS_154 (specific to CoLk94184) and mSSCIR43_238 (specific to CoSe92423)] in sugarcane breeding, vis a vis transporting CE-based evaluation in clone/variety identity testing, cross fidelity assessments, and genetic relatedness among Saccharum species and related genera.