Location: Subtropical Horticulture ResearchTitle: Pooled-BAC sequencing of a black pod resistance region (cBPQTL12) in T. cacao Author
|Schnell Ii, Raymond|
Submitted to: Annual International Plant & Animal Genome Conference
Publication Type: Abstract Only
Publication Acceptance Date: 1/16/2011
Publication Date: 1/16/2011
Citation: Feltus, F.A., Saski, C.A., Mockaitis, K., Choi, J., Blackmon, B., Ficklin, S.P., Haiminen, N., Kuhn, D.N., Motamayor, J.C., Schnell II, R.J. 2011. Pooled-BAC sequencing of a black pod resistance region (cBPQTL12) in T. cacao. Annual International Plant & Animal Genome Conference. 2011. Interpretive Summary: Theobroma cacao, the source of cocoa beans for chocolate, is an important tropical agriculture commodity that is affected by a number of fungal pathogens and insect pests, as well as concerns about yield and quality. We are trying to find molecular genetic markers that are linked to disease resistance and other important economic traits to aid in a marker assisted selection (MAS) breeding program for cacao to ensure a reliable supply of cocoa for the US confectionary industry. Currently there are about 500 molecular genetic markers for cacao and we are taking advantage of the cacao genome sequencing project to expand that to greater than 50,000 single-nucleotide polymorphism (SNP) markers. Here, we sequence a 3 Mb region of the genome known to be associated with black pod resistance to identify candidate genes associated with resistance to use as markers for MAS. The assembly of the genome sequence is a critical step in the resulting identification of markers for the mapped traits. Our results are important to scientists trying to understand the mechanism of disease resistance and, eventually, to cacao farmers who will benefit from superior disease resistant and more productive cultivars produced through our MAS breeding program.
Technical Abstract: Whole genome sequencing (WGS) is an expensive and technically challenging endeavor. An alternative to WGS is to sequence specific chromosomal segments of biological interest (e.g. a QTL interval). This method is cheaper than WGS and reduces the risk of misassembly from distal parts of the genome. Using the minimum tiling path (MTP) from the T. cacao physical map, we have identified a ~3Mbp region (27 BACs) that corresponds with a black pod resistance QTL (cBPQTL12). BAC clones from this section of the MTP were isolated, bar-coded, 454/Roche sequenced as a BAC pool, and assembled at various mixtures of linear and paired end reads. In addition, a sub-region containing 10 BACs was sequenced using the traditional Sanger technique for assembly accuracy comparison. The practical aspects of linear/paired read mixture and depth ratio selection will be presented. Annotation of the region has identified candidate genes that could be controlling the black pod resistance QTL.