Genetic analysis of the Group IV Rht LOCI in wheat.
The introduction of the group IV semi-dwarf Rht alleles, Rht-B1b (formerly Rht1) and Rht-D1b (formerly Rht2) into bread wheat varieties from the donor line „Norin 10‟ that began in the 1960s was a major contributor to the „green revolution‟. Rht-B1b and Rht-D1b were characterised and cloned over a decade ago (Gale and Youssefian 1985; Peng et al. 1999), however the Rht-A1 locus has not been isolated and little is known regarding the genetic diversity of the group IV Rht loci or the genetic composition of the contiguous sequence surrounding Rht that was presumably introgressed into wheat varieties along with the dwarfing alleles. To investigate the contiguous region around Rht, a hexaploid wheat („Chinese Spring‟ (CS)) BAC library was screened using a PCR-based technique (Febrer et al. 2009). This identified several Rht-containing BAC clones, three of which (representing the A, B, and D genomes) were sequenced and found to contain one to two genes upstream of Rht in conserved order. Gene synteny was also highly conserved in rice, Brachypodium distachyon, sorghum, and maize. The previously unidentified Rht-A1 homoeologue was physically mapped to the long arm of chromosome 4A using aneuploid lines and mapped relative to genetic markers. To estimate genetic diversity, the entire coding regions of Rht-A1, Rht-B1, and Rht-D1 and the flanking regions (approximately 1800 bp 5‟ and 450 bp 3‟) were sequenced in 40 diverse wheat accessions. Little polymorphism and few haplotypes were identified on the A and D genomes, but on the B genome a relative abundance of haplotypes and polymorphism were present, including insertions (relative to CS) of 160 bp and 197 bp within 600 nucleotides of the ORF. The Rht-B1 insertions did not have a pronounced effect on RNA transcript level when assessed in seedlings. In an analysis of 368 lines from the INRA bread wheat core collection (BWCC) (Balfourier et al., 2007), lines with the Rht-B1 insertions were associated with reduced heights and reduced GA sensitivity relative to lines without an insertion, but only the height reductions associated with the 197 bp insertion were significant (p < 0.05). GA sensitivity tests of the INRA BWCC did not reveal any novel GA insensitive mutants. An investigation of the origin of the „Norin 10‟ alleles revealed potential discrepancies between the published pedigree and of „Norin 10‟ and the genotypes of seed stocks. Sequence, annotation, and comparative genomics of the Rht-containing BAC clones, the mapping of Rht-A1, and the discovery and investigation of novel genetic diversity provides greater insight into the Rht region and also provides tools for further analysis of this region and for the potential improvement of bread wheat. *Article from 'Genome' submitted as an appendix (further bibliographic details contained within thesis contents) not uploaded. Please contact author for further information. Article available on submitted CD deposited with UEA Library.
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