Screening the R570 sugarcane BAC library to identify BAC clones that underlie QTL of agronomic importance

Sugarcane

In collaboration with:

Dr Karen Aitken

CSIRO Plant Industry

Queensland Bioscience Precinct

306 Carmody Road                                                                     

St Lucia

4068

QLD Australia

Email: Karen.Aitken@csiro.au

Abstract:

Over the last decade a number of genetic maps have been constructed and quantitative trait loci (QTL) studies carried out for agronomically important traits. These have included traits related to sugar content (Brix, Pol), as well as yield and biomass related traits such as stem diameter, height and stalk number. In parallel a large association study using the parental population of the Australian sugarcane breeding program has also been conducted targeting disease resistance. These studies have identified many markers that contribute to the phenotypic variation of these traits. The next step is to determine the genetic structure of these regions, and which genes or regulatory elements underlie these QTL. In the last five years a consortium of scientists from a number of countries including Australia have grouped together to form the Sugarcane Genome Sequencing Initiative (SUGESI). The goal of this consortium is to generate a combined monoploid genome sequence of sugarcane. The Australian contribution to this consortium is sequenced BAC clones from the R570 BAC library that underlie the QTLs identified in the numerous studies. The R570 BAC library will be screened at INRA-CNRGV to identify the BAC clones that underlie these QTL of agronomic importance.

The aims of this project are:

1) To establish a strategy to identify which QTL regions to target.

QTL regions were selected that had been identified at a significance level of less than p=0.001 and contained more than one marker. Ideally, the selected QTL should also have been identified in more than one population. In the next stage, sets of linked markers were selected to test across the whole QTL region. The selected markers were in most cases SNP or DArT markers with a known sequence. The sequenced markers were BLASTn aligned to the sorghum genome to determine the abundance of the sequence. Only those markers that aligned to a single location in sorghum were selected to screen the BAC clones and preference was given to those that also showed alignment to the sugarcane EST collection.

2) Screen the R570 BAC library with the selected markers linked to QTL.

Two different approaches: macroarray membrane hybridisation and qPCR using 3-D BAC pools will be used to screen the library. In total 112 primer pairs were developed to screen the R570 BAC library. These will be used to identify BAC clones which will then be characterised and ultimately sequenced to identify genes in these regions. Once the sequence is obtained a major outcome will be the development of robust markers, closely linked to causal genes, for use in the Australian sugarcane breeding program.