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Purification of High Molecular Weight Genomic DNA from Powdery Mildew for Long-Read Sequencing
Published on: March 31, 2017
Molecular identification of a Pm4 allele conferring powdery mildew resistance in durum wheat DR88
Guohao Han1, Lixian Xing1, Tiantian Gu1
1Center for Agricultural Resources Research, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Shijiazhuang, 050022, China.
Background:
Powdery mildew, caused by Blumeria graminis f. sp. tritici (Bgt), is one of the most destructive wheat diseases worldwide. Durum wheat (Triticum turgidum L. var. durum Desf.) is a crucial gene donor for improving common wheat.
Results:
In this study, we investigated a durum wheat accession, DR88, which exhibits broad and high levels of resistance to powdery mildew. Using bulked segregant RNA-Seq (BSR-Seq), we identified a dominant gene, tentatively designated PmDR88, and localized it to 743-776 Mb interval on chromosome arm 2AL according to the reference genome of durum wheat cv. Svevo. Subsequently, PmDR88 was mapped in a genetic region of 3.9 cM flanked by the markers WGRE77410 and WGRC872 at genetic distances of 1.6 and 2.3 cM, respectively; it also co-segregated with JS717×JS718, the diagnostic marker for the Pm4 locus. Genotyping of a large population comprising 5,174 F2:3 families using JS717×JS718 confirmed that PmDR88 is located at the Pm4 locus on 2AL. Sequence alignment revealed that PmDR88 shares identical amino acid sequences with Pm4d, while qRT-PCR analysis suggested distinct expression patterns for PmDR88 compared with previously reported Pm4 alleles. Two complementary DNA markers, including the dominant co-segregating marker JS717×JS718 and a newly developed closely-linked co-dominant marker WGRE77410, were confirmed to be available for efficiently transferring PmDR88 into the tested wheat backgrounds by marker-assisted selection (MAS) strategy.
Conclusions:
PmDR88 was mapped in the Pm4 locus. Despite sharing identical amino acid sequences with Pm4d, PmDR88 exhibits distinct expression patterns. Moreover, DR88 shows broad and high levels of resistance to powdery mildew. Two complementary DNA markers were identified for MAS breeding. The molecular identification of PmDR88 will facilitate transfer of this Pm4 allele into susceptible cultivars for resistance improvement or into resistant cultivars for resistance-enhanced pyramiding breeding.
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