Efficient development of Haynaldia villosa chromosome 6VS-specific DNA markers using a CISP-IS strategy

Development of effective molecular markers linked to Pm21 deriving from Haynaldia villosa is critical for wheat breeding of powdery mildew resistance. In this study, we designed 12 pairs of conserved-intron scanning primers (CISPs), using intron-containing conserved genes located on the short arm of Brachypodium distachyon chromosome 3 (3BdS) aligned with cDNA or expressed sequence tags (ESTs) of Triticeae crops. Of 12 CISP primer pairs, 11 amplified DNA both in H.villosa and in wheat, and four displayed H.villosa chromosome 6VS-specific polymorphisms. Six non-polymorphic DNAs were further sequenced for designing internal primers, and five additional 6VS-specific markers were obtained. Of the total nine 6VS-specific co-dominant markers, six could effectively trace Pm21 in F2 population derived from the hybrid between the T6AL.6VS line and Yangmai 158'. This study demonstrated that Brachypodium genomic information could be powerfully utilized to develop molecular markers in H.villosa or other Triticeae species.