Identification and mapping of conserved ortholog set (COS) II sequences of cacao and their conversion to SNP markers for marker-assisted selection in Theobroma cacao and comparative genomics studies

Abstract

Theobroma cacao (cacao) is a tree cultivated in the tropics around the world for its seeds that are the source of both chocolate and cocoa butter. Genetic marker development for marker-assisted selection (MAS) is critical for the success of cacao breeding for disease resistance and yield. To develop conserved ortholog set II (COSII) singlenucleotide polymorphism (SNP) markers for MAS in cacao, we have used three strategies and three types of cacao genetic and sequence data to identify and map 98 cacao COSII genes. The resources available at the time these studies were first undertaken dictated the strategy utilized. For the first strategy, SNPs were identified using cacao expressed sequence tags homologous to COSII sequences. Strategy II utilized a leaf transcriptome of cacao genotype "Matina 1-6" and Strategy III the genomic sequence of a 3-Mb region of "Matina 1-6" linkage group 5 associated with an important quantitative trait locus (QTL) for resistance to black pod. We have identified SNP markers for 83 of the 98 mapped COSII genes, and 19 of these SNP markers co-locate with QTLs. These COSII SNP markers, the first identified for cacao, will be used for genotyping and off-typing in cacao breeding programs and employed for genetic mapping and syntenic studies to trace co-location of genes regulating traits of importance between cacao and other species.

Making a chocolate chip: development and evaluation of a 6K SNP array forTheobroma cacao

“…This transcriptome represented ∼22 Mb of sequence in 29,787 isotigs. 40 Paired-end Illumina GAII reads for 16 highly diverse cacao accessions were obtained and mapped as described (SRA BioProject PRJNA51633). Supplementary Fig.…”

“…Total leaf RNA of Matina 1-6 (800 ng) was used for a library preparation optimized for Roche/454 GS FLX Titanium sequencing described previously (NCBI SRX172965 24 , 40 ). High-quality sequence reads (1,510,557) were trimmed and assembled using Newbler v2.0 into 29,787 contigs of average length 725 (SD = 558).…”

Identification of candidate genes involved in Witches’ broom disease resistance in a segregating mapping population of Theobroma cacao L. in Brazil

“…DNA for genotyping was isolated from leaf samples according to the protocol described by Motamayor et al [ 6 ]. The SNPs that were used on the cacao Illumina Infinium SNP chip were identified as part of the work to identify conserved ortholog set II (COSII) SNP markers for MAS in cacao as described by Kuhn et al [ 21 ] and Livingstone et al [ 22 ]. The strategy was to sequence leaf RNA samples of 15 members of a panel of diverse cacao genotypes and to align the sequences against the Theobroma cacao Matina 1–6 leaf transcriptome.…”

Cacao biotechnology: current status and future prospects

“…In this context, DNA fingerprinting as a screening tool has been extensively used in rapid and accurate identification of cacao accessions. Restriction fragment length polymorphisms (RFLPs), random amplified polymorphic DNA (RAPD), amplified fragment length polymorphisms (AFLPs), microsatellites and single nucleotide polymorphisms (SNPs) are some of the molecular markers commonly used in cacao molecular studies (Kuhn et al ., ; Lanaud et al ., ; Laurent et al ., ; Motilal and Butler, ; Santos et al ., ; Turnbull et al ., ). Livingstone et al .…”