Mapping and Tagging of Simply Inherited Traits - Genetics, Genomics and Breeding of Crop Plants - page 123

( Pl 6 ), HA338 ( Pl 7 ), RHA340 ( Pl 8 ), YDQ ( Pl 6 ), XRQ ( Pl 5 ), and susceptible line],
and RHA419 that obtained its downy mildew resistance from H. argophyllus
(Arg1575-2). Their results indicate that RHA419 carries a single gene or a
cluster of resistance genes to many downy mildew races and is independent
of the Pl 6 / Pl 7 and Pl 5 / Pl 8 clusters. Dussle et al. (2004) confirmed this finding
by identifying 12 polymorphic SSRs with BSA in the F 2 progenies of the
cross cmsHA342 x Arg1575-2 ( Pl ARG ). The 12 SSR markers and Pl ARG were
mapped on LG 1.
A large number of resistance ( R ) genes from different plant species were
cloned and analyzed in the last few years (Gebhardt 1997; Michelmore
2000). The comparison of the sequence revealed that R -genes contain
conserved domains. Two groups of NBS-LRR (nucleotide-binding site—
leucine-rich repeats) resistance gene candidates have been described in
plants: the TIR-NBS-LRR subclass (the TIR domain is homologous to the
Drosophila Toll and mammalian Interleukin-1 receptor) and the non-TIR-
NBS-LRR subclass. The resistance gene analog (RGA, also termed resistance
gene candidate—RGC) approach allows the search of new Pl resistance
genes in wild sunflower species. It also enables the analysis of resistance
mechanisms against P. halstedii and the development of molecular markers
derived from the gene itself or from closely linked sequences as a starting
point for map-based cloning or MAS.
Gentzbittel et al. (1998) first used an RGA approach to analyze the
resistance of sunflower to downy mildew. Degenerate primers were designed
from the conserved NBS domain of the virus resistance gene N from tobacco
(Whitham et al. 1994) and the rust resistance gene L6 from flax (Lawrence et
al. 1995). After cloning and sequencing, one RGA (NBS-R3), belonging to
the TIR-NBS-LRR group, was mapped in three populations in the region of
the Pl loci on group 1 of the CARTISOL map (Gentzbittel et al. 1995).
Bouzidi et al. (2002) conducted further analysis on the complexity of
the Pl 6 locus in order to develop molecular markers suitable for positional
cloning of Pl genes. To clone a full-length cDNA RGA, the sequence of the
RGA product HA-NBS3 (Gentzbittel et al. 1998) was used as a template in
RACE-PCR to obtain 5' and 3' ends of the cDNA. By using the sequence of
the full-length RGA (GenBANK accession number AF316405) specific
primers were designed and tested for polymorphism using BSA. Thirteen
dominant sequence tagged site (STS) markers were obtained covering a
genetic distance of about 3 cM centered on the Pl 6 locus. All 13 STS markers
showed homology to the TIR-NBS-LRR subclass. Pankovic et al. (2007)
deployed the three specific primer pairs HAP1, HAP2, HAP3 of Bouzidi et
al. (2002) for screening a near-isogenic line (NIL) population [HA26 S x NIL
HA26 R ( Pl 6 )]. The fragment HAP3/S1 (resembling Ha-NBS 12S; Bouzidi et
al. 2002) was the only one, which was observed for both resistant and
susceptible parental line. Based on this RGA, two co-dominant cleaved