Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 12/15/2006
Publication Date: 3/20/2007
Citation: Liu, Z., Faris, J.D., Edwards, M.C., Friesen, T.L. 2006. The initiation of map-based cloning of an avirulence gene from Pyrenophora teres. Meeting Abstract. 24th Fungal Genetics Conference. p. 159.
Technical Abstract: The fungus Pyrenophora teres, causal agent of barley net blotch, is an economically important pathogen in most barley-growing areas. In a previous study, we identified and mapped a fungal gene associated with avirulence on Tifang barley that was already designated AvrHar. This study used a P. teres mapping population consisting of parental isolates 15A (avirulent on Tifang) and 0-1 (virulent on Tifang). In the present study, an attempt is being made to clone AvrHar using a map-based approach. Initially, the fungal population used in the mapping of AvrHar was enlarged from 78 progeny to a total of 278 progeny to create a map with increased resolution. A single AFLP marker closely linked to the Avr gene was identified and TAIL-PCR was used to extend the fragment from 250bp to 1.7kb for use as a probe. A 20X BAC library was constructed from parental isolate 0-1 and a 3X fosmid library was constructed from parental isolate 15A. The AFLP probe allowed the identification of 14 positive clones from the 0-1 BAC library. A single BAC clone was sequenced and shown to contain a large portion of Gypsy-like LTR-retrotransposon repeats at both ends. A 60kb region in the middle of the BAC contained several predicted genes. A PCR marker based on one of the genes located approximately 50kb from the original AFLP was found to co-segregate with the AFLP marker, indicating that AvrHar has not been reached. An additional 4 BAC clones have been identified using the new marker. Twelve fosmid clones spanning approximately 90 kb outside the BAC clone sequence have been characterized. Recombination in this region is lower than expected and several walking steps may be necessary. Additional steps are being taken to span AvrHar.