Cytogenetics and genome structure in genus Miscanthus, a potential source of bioenergy feedstocks

Chae, Won Byoung

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https://hdl.handle.net/2142/34308 Copy

Description

Title

Cytogenetics and genome structure in genus Miscanthus, a potential source of bioenergy feedstocks

Author(s)

Chae, Won Byoung

Issue Date

2012-09-18T21:10:36Z

Director of Research (if dissertation) or Advisor (if thesis)

Juvik, John A.

Doctoral Committee Chair(s)

Juvik, John A.

Committee Member(s)

• Moose, Stephen P.
• Rayburn, A.L.
• Ming, Ray R.
• Sacks, Erik J.

Department of Study

Crop Sciences

Discipline

Crop Sciences

Degree Granting Institution

University of Illinois at Urbana-Champaign

Degree Name

Ph.D.

Degree Level

Dissertation

Keyword(s)

• Miscanthus
• Genome size
• genetic mapping
• genome doubling

Abstract

Miscanthus (subtribe Saccharinae, tribe Andropogoneae, family Poaceae) is a genus of temperate perennial C4 grasses. Accessions in the genus Miscanthus are potential crop candidates of lignocellulosic biomass for energy production, whose high biomass productivity is attractive as a biofuel feedstock. Miscanthus is not native to the United States and has been introduced as ornamental accessions by private nurseries and research institutes. Comprehensive taxonomic studes have not been conducted on these U.S. accessions. Previous taxonomic studies have been conducted on the genus Miscanthus using morphology and DNA sequence variation. This study (chaper 2) combines information on genome size and ploidy levels and DNA sequence variation to classify Miscanthus accessions to aid potential biomass crop improvement programs and to investigate the evolution of the genus. We observed that Miscanthus accessions fell into 4 groups, including section Miscanthus, section Triarrhena and two groups intermediate between two sections, based on morphology and genome size. Sixteen simple sequence repeat (SSR) primer pairs were selected based on amplification and polymorphism across three genera, Miscanthus, Saccharum and Erianthus. Morphology, genome size and SSR genotyping of 42 accessions including diploid and triploid interspecific hybrid progeny suggested that there are three Miscanthus species (M. sinensis, M. sacchariflorus and M. x giganteus), one M. sacchariflorus variety and one putative hybrid among Miscanthus accessions, which were clearly separated from the other two genera. The species status of M. floridulus remains in question. The evolution of Miscanthus and related genera is discussed based on genome size, ploidy level, cluster analysis and geographical distribution. Based on genome size and chromosome number comparision between Miscanthus and other related genera, we hypothesize large-scale duplications have occurred in recent ancestors of Miscanthus. Owing to the complexity of the Miscanthus genome and the complications of self-incompatibility, a complete genetic map with a high density of markers has not yet been developed. As described in chapter 3, a cross between two M. sinensis accessions, ‘Grosse fontaine’ and ‘Undine’ was made to produce 221 segrating progeny as a mapping population. Simple sequence repeat (SSR) markers from sugarcane expressed sequence tags (EST) and genomic sequences were screened in the two parental M. sinensis accessions. Single nucleotide polymorphism (SNP) markers from deep transcriptome sequencing (RNAseq) were also used for map construction. A total of 210 SSR markers and 658 single nucleotide polymorphism (SNP) markers were validated via segregation in the full sib F1 mapping population. A genetic map for M. sinensis was constructed that was resolved into 19 linkage groups, the haploid chromosome number expected from cytological evidence. Comparative genomic analysis revealed genome-wide duplication in Miscanthus relative to S. bicolor, with subsequent insertional fusion of a pair of chromosomes. The utility of the map is confirmed by the identification of two paralogous C4-pyruvate, phosphate dikinase (C4-PPDK) loci in Miscanthus, at positions syntenic to the single orthologous gene in sorghum. The M. sinensis map and comparative mapping with sorghum suggests that the genus Miscanthus experienced an ancestral tetraploidy and chromosome fusion prior to its diversification, but after its divergence from the closely related sugarcane clade. The genetic map for Miscanthus is useful in biological discovery and breeding efforts to improve this emerging biofuel crop, and also provide a valuable resource for understanding genomic responses to tetraploidy and chromosome fusion. In chapter 4, artificial genome doubling of various Miscanthus accessions with antimototic agents was used to understand the phenotypic responses to whole genome duplication in Miscanthus. Interspecific manipulation of ploidy levels is also a potential strategy for Miscanthus crop improvement to generate superior germplasm and to circumvent reproductive barriers for the introduction of new genetic variation into core germplasm. Therefore, synthetic autotetraploid lines of M. sacchariflorus and M. sinensis, and autoallohexaploid M. x giganteus were produced in tissue culture from oryzalin treatments to seed- and immature inflorescence-derived callus lines. Genome doubling of diploid M. sinensis, M. sacchariflorus, and triploid M. x giganteus to generate tetraploid and hexaploid lines was confirmed by stomata size, nuclear DNA content, and chromosome counts. A putative pentaploid line was also identified among the M. x giganteus synthetic polyploid lines by nuclear DNA content and chromosome counts. Comparisons of phenotypic performance of synthetic polyploid lines with their diploid and triploid progenitors in the greenhouse found species-specific differences in plant tiller number, height, and flowering time among the doubled lines. Stem diameter tended to increase after polyploidization but there were no significant improvement in biomass traits. Under field conditions, M. x giganteus synthetic polyploid lines showed greater phenotypic variation, in terms of plant height, stem diameter and tiller number, than their progenitor lines. Production of synthetic autopolyploid lines displaying significant phenotypic variation suggests that ploidy manipulation can introduce genetic diversity in the limited Miscanthus germplasm currently available in the United States. The role of polyploidization in the evolution and breeding of the genus Miscanthus is discussed.

Graduation Semester

2012-08

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Copyright 2012 Won Byoung Chae

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