title: High-throughput 454 resequencing for allele discovery and recombination mapping in Plasmodium falciparum.
abstract: BACKGROUND
  Knowledge of the origins, distribution, and inheritance of variation in the malaria parasite (Plasmodium falciparum) genome is crucial for understanding its evolution; however the 81% (A+T) genome poses challenges to high-throughput sequencing technologies. We explore the viability of the Roche 454 Genome Sequencer FLX (GS FLX) high throughput sequencing technology for both whole genome sequencing and fine-resolution characterization of genetic exchange in malaria parasites.
  
  
  RESULTS
  We present a scheme to survey recombination in the haploid stage genomes of two sibling parasite clones, using whole genome pyrosequencing that includes a sliding window approach to predict recombination breakpoints. Whole genome shotgun (WGS) sequencing generated approximately 2 million reads, with an average read length of approximately 300 bp. De novo assembly using a combination of WGS and 3 kb paired end libraries resulted in contigs ≤ 34 kb. More than 8,000 of the 24,599 SNP markers identified between parents were genotyped in the progeny, resulting in a marker density of approximately 1 marker/3.3 kb and allowing for the detection of previously unrecognized crossovers (COs) and many non crossover (NCO) gene conversions throughout the genome.
  
  
  CONCLUSIONS
  By sequencing the 23 Mb genomes of two haploid progeny clones derived from a genetic cross at more than 30× coverage, we captured high resolution information on COs, NCOs and genetic variation within the progeny genomes. This study is the first to resequence progeny clones to examine fine structure of COs and NCOs in malaria parasites.
authors.affiliation: Department of Biological Sciences, Eck Institute for Global Health, University of Notre Dame, Notre Dame, IN 46556, USA.
authors.forename: Upeka
authors.initials: U
authors.lastname: Samarakoon
authors.forename: Allison
authors.initials: A
authors.lastname: Regier
authors.forename: Asako
authors.initials: A
authors.lastname: Tan
authors.forename: Brian A
authors.initials: BA
authors.lastname: Desany
authors.forename: Brendan
authors.initials: B
authors.lastname: Collins
authors.forename: John C
authors.initials: JC
authors.lastname: Tan
authors.forename: Scott J
authors.initials: SJ
authors.lastname: Emrich
authors.forename: Michael T
authors.initials: MT
authors.lastname: Ferdig
mesh_terms: D000483:Alleles; D056905:Chromosome Breakpoints; D002874:Chromosome Mapping; D003433:Crosses, Genetic; D016054:DNA, Protozoan / Q000235:genetics; D005785:Gene Conversion; D018628:Gene Dosage; D018503:Genome, Protozoan; D015698:Genomic Library; D005838:Genotype; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D018895:Microsatellite Repeats; D010963:Plasmodium falciparum / Q000235:genetics*; D020641:Polymorphism, Single Nucleotide; D017422:Sequence Analysis, DNA / Q000379:methods
chemical_list: D016054:DNA, Protozoan