Open Access Highly Accessed Methodology article

High-throughput 454 resequencing for allele discovery and recombination mapping in Plasmodium falciparum

Upeka Samarakoon1, Allison Regier12, Asako Tan1, Brian A Desany3, Brendan Collins1, John C Tan1, Scott J Emrich12 and Michael T Ferdig1*

Author Affiliations

1 Department of Biological Sciences, Eck Institute for Global Health, University of Notre Dame, Notre Dame, IN 46556, USA

2 Department of Computer Science and Engineering, University of Notre Dame, Notre Dame, IN 46556, USA

3 454 Life Sciences, a Roche company, 15 Commercial Street, Branford, CT 06405, USA

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BMC Genomics 2011, 12:116  doi:10.1186/1471-2164-12-116

Published: 17 February 2011

Additional files

Additional file 1:

(A+T) content of WGS reads in uniquely mapped regions of parents and progeny

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Additional file 2:

SNPs between HB3 and Dd2

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Additional file 3:

Refined views of previously unknown COs shown in Figure 3. Previously unknown COs detected in the progeny lines (highlighted with black bars in Figure 3) are indicated by double arrows in the chromosomal view (top) and zoomed-in view (boxed, bottom). (A-D) previously unknown COs in 7C126, (E-H) previously unknown COs in SC05. SNP map by 454 sequencing is presented in comparison with the MS marker linkage map in P. falciparum [4]. Each line represents a single SNP marker. HB3 alleles are shown in green bars and Dd2 alleles are shown in red.

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Additional file 4:

Visual inspection of mapped WGS sequence at CO and NCO gene conversion breakpoints shown in Figure 4. (A) simple CO breakpoint in 7C126, (B) NCO gene conversion in SC05 and (C) complex CO breakpoint accompanied by a conversion tract (defined by rapid allele changes at/near breakpoint) detected in 7C126. Chromosomal alignments at CO and NCO regions were visually inspected in comparison with the parental genomes, using Integrative Genomics Viewer [45]. (i) SNPs detected by 454 sequencing are ordered according to the chromosomal location (PlasmoDB v5.4 [27]). Each line represents a single SNP marker. HB3 alleles are shown in green bars and Dd2 alleles are shown in red. (ii) Comparison of SNPS and read alignments at selected SNP loci (arrow). SNPs are highlighted in blue [C], red [T], green [A] and brown [G].

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Additional file 5:

Resequencing results of de novo SNP positions

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Additional file 6:

Distance between consecutive de novo SNPs. The distance between consecutive de novo SNPs were calculated to detect SNP clustering characteristic of sequencing errors or mis-mapping errors. 35% of the de novo SNPs were clustered in distances of less than 5 bps.

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Additional file 7:

Base conversion trends in de novo SNPs. The type of base conversion was investigated for positions at which the parental base calls were identical. More transversions were detected for SC05 compared to 7C126 (A), but did not show a predominant base conversion bias from 7C126 (B).

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Additional file 8:

Alternate SNP positions. The alternate SNP positions were assessed for their primary and secondary positions base call identity. Most primary base calls reflected the parental base call. The secondary base call position varied in the 2 progeny genomes in base call identity. Majority of the secondary base calls were parental in 7C126, whereas majority of the secondary base calls were non-parental in SC05 (A). Majority of the primary base calls were transitions in 7C126, while they were transversions in SC05 (B, C).

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Additional file 9:

Selected CNVs in 7C126 and SC05. Comparative genomic hybridization (CGH) was used to detect large (> 1 kb) CNV regions in 7C126 (A) and SC05 (B). Five known CNVs that exist between parental strains were detected in the progeny in Chr 2, 5, 9 and 12.

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