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Developing High Throughput Genotyped Chromosome Segment Substitution Lines Based on Population Whole-Genome Re-Sequencing in Rice (Oryza Sativa L.)

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Developing High Throughput Genotyped Chromosome Segment Substitution Lines Based on Population Whole-Genome Re-Sequencing in Rice (Oryza Sativa L.)

Jianjun Xu et al. BMC Genomics.

Abstract

Background: Genetic populations provide the basis for a wide range of genetic and genomic studies and have been widely used in genetic mapping, gene discovery and genomics-assisted breeding. Chromosome segment substitution lines (CSSLs) are the most powerful tools for the detection and precise mapping of quantitative trait loci (QTLs), for the analysis of complex traits in plant molecular genetics.

Results: In this study, a wide population consisting of 128 CSSLs was developed, derived from the crossing and back-crossing of two sequenced rice cultivars: 9311, an elite indica cultivar as the recipient and Nipponbare, a japonica cultivar as the donor. First, a physical map of the 128 CSSLs was constructed on the basis of estimates of the lengths and locations of the substituted chromosome segments using 254 PCR-based molecular markers. From this map, the total size of the 142 substituted segments in the population was 882.2 Mb, was 2.37 times that of the rice genome. Second, every CSSL underwent high-throughput genotyping by whole-genome re-sequencing with a 0.13× genome sequence, and an ultrahigh-quality physical map was constructed. This sequencing-based physical map indicated that 117 new segments were detected; almost all were shorter than 3 Mb and were not apparent in the molecular marker map. Furthermore, relative to the molecular marker-based map, the sequencing-based map yielded more precise recombination breakpoint determination and greater accuracy of the lengths of the substituted segments, and provided more accurate background information. Third, using the 128 CSSLs combined with the bin-map converted from the sequencing-based physical map, a multiple linear regression QTL analysis mapped nine QTLs, which explained 89.50% of the phenotypic variance for culm length. A large-effect QTL was located in a 791,655 bp region that contained the rice 'green revolution' gene.

Conclusions: The present results demonstrated that high throughput genotyped CSSLs combine the advantages of an ultrahigh-quality physical map with high mapping accuracy, thus being of great potential value for gene discovery and genetic mapping. These CSSLs may provide powerful tools for future whole genome large-scale gene discovery in rice and offer foundations enabling the development of superior rice varieties.

Figures

Figure 1
Figure 1
The locations of the polymorphic markers in the rice physical map.
Figure 2
Figure 2
Flowchart of the development of CSSLs in the present study. MAS: marker-assisted selection.
Figure 3
Figure 3
The physical map and bin-map of the 128 chromosome segment substitution lines (CSSLs). (A) The physical map of the CSSLs was constructed with molecular markers. Each row represented a CSSL and each column represented a molecular marker locus. The black areas indicate regions that were homozygous for Nipponbare alleles; the white areas indicate regions homozygous for 9311 alleles. (B) The physical map of the CSSLs constructed by whole-genome resequencing. The blue areas indicate regions that are homozygous for Nipponbare alleles; the white areas indicate regions that are homozygous for 9311 alleles. (C) Bin-map of the CSSLs. The blue areas indicate regions that are homozygous for Nipponbare alleles; the red areas indicate regions homozygous for 9311 alleles.
Figure 4
Figure 4
Distribution of the length of the substituted chromosome segments in the 128 CSSLs. MM-map: based on the physical map constructed with molecular markers; GR-map: based on the physical map constructed by whole-genome resequencing.
Figure 5
Figure 5
CL of CSSLs under natural field conditions. (a) The CL phenotypes in 9311 and CSSLs. Scale bar, 50 cm. (b) Distribution of CLs in 128 CSSLs under natural field conditions.
Figure 6
Figure 6
Recombination map of CSSL 89. Red lines: homozygous 9311 genotype; blue lines: homozygous Nipponbare genotype; green panes: double-crossovers.
Figure 7
Figure 7
A hypothetical CSSL library with eight CSSLs, a donor parent and a recurrent parent. The red areas indicate regions that are homozygous for donor alleles; the green areas indicate regions homozygous for recurrent alleles.

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