No description available.

Arabidopsis expressed sequence tags (ESTs) and microsatellites of Brassica napus have been developed and used as PCR-based markers for both mapping and genetic diversity studies in B. napus . Out of 300 random Arabidopsis ESTs screened, 43 markers were mapped onto a genetic map of B. napus and then used in a diversity study involving 48 B. napus cultivars. A second set of EST markers were developed from chromosome 1 of Arabidopsis and used in genetic mapping studies of B. napus. From 192 primer pairs developed, 50 markers were added onto the B. napus reference map. Microsatellite markers were developed using a "GA" enriched genomic library from B. napus. From 152 designed primer pairs, 23 markers were added onto the B. napus reference map. Microsatellite markers were also used in genetic diversity studies of B. napus, where, from the 152 primer pairs, 40 revealed polymorphism between the 48 B. napus cultivars.

Frost can be detrimental to canola (Brassica napus L.) production. Depending on the severity, the entire field can be killed. Having frost tolerance in canola would benefit growers by allowing them to plant early, utilize early season moisture, and avoid high heat during flowering. However, frost tolerance in canola has not been well studied. A protocol was developed that determined 14 day old seedlings should be acclimated at 4°C for 7 days before being exposed to overnight frost (-4°C) in a small freezing chamber. However, when a larger chamber was used for freezing, the protocol was optimized to -8°C instead. A greenhouse study was conducted on a diverse collection of 231 genotypes and genome-wide association scan (GWAS) was conducted to identify potential genes that were related to frost tolerance or abiotic stress tolerance. Thirty-eight significant single nucleotide polymorphism (SNP) markers were selected based on 10,000 bootstraps and 0.1 percent tail of the empirical distribution. The markers were located on chromosomes A01, A02, A03, A04, A07, A08, A09, A10, C03, C05, C06, C07, and C09. Stepwise regression highlighted a QTL located on chromosomes A02. Another GWAS was done on 147 canola germplasm lines phenotyped under natural conditions. Thirty-eight significant SNPs identified from this study were located on chromosomes A05, A07, A09, C01, C02, C03, C04, C05, C06, C07, and C09. Stepwise regression identified a QTL located on chromosome C04. A protocol was developed to measure the freezing induced electrolyte leakage from leaves of rapeseed/canola. A total of 157 germplasm lines were evaluated for freezing induced (-12°C for 2 h) electrolyte leakage. Thirty-six significant SNPs located on chromosomes A01, A02, A03, A04, A05, A06, A07, A08, A09, A10, C01, C02, C04, C05, C06, C07, and C09 were identified. Stepwise regression identified 10 QTL located on chromosomes A01, A02, A04, A06, A07, C02, C05, C07, C09, and one that could not be assigned. All GWAS studies identified potential genes of interest that were related to frost tolerance, abiotic stress, and transcription factors. / Northern Canola Growers Association

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Bibliography: leaves 132-159. Boron efficiency in oilseed rape (Brassica napua L. and B. juncea L.) was investigated in a wide range of genotypes. Using a solution culture screening of 10 day old seedlings, root length best described shoot growth response, and was used to characterise a total of 65 genotypes. Varieties and breeders lines tolerant of B-deficient growing conditions were identified, and the screening process validated through field trials. B responses in plants sampled at the 'green bud' stage indicated that vegetative growth is important in B efficiency. Studies were conducted to investigate the mechanism of B efficiency in oilseed rape. Results suggest no association between B efficiency and the capacity to acidify the root rhizosphere, or an increased translocation of B from root to shoot. Boron retranslocation was also studied as a mechanism of B efficiency.

Triacylglycerol (TAG) production in the microspore derived (MD) cell suspension culture ofBrassica napus L. cv Jet Neuf was enhanced when the sucrose concentration in the growth medium was increased from 2 to 14 % (w/v). mRNA differential display by polymerase chain reaction was used to examine gene expression in cells grown at different sucrose concentrations in order to identify mRNAs which could be associated with oil formation. The anchored primer, T12AA, was used to screen one subset, representing approximately one twelfth of the transcript population, isolated from cultures grown in media supplemented to 2, 6 and 14 % (w/v) sucrose. Analysis of this mRNA subset revealed thirteen cDNAs which appeared to be upregulated as the sucrose concentration was increased. Cloning and sequencing revealed multiple cDNA fragments for each signal detected by differential display. RT-PCR analysis of sixteen different cDNAs revealed that eight encoded mRNAs which were upregulated in parallel to the increase in media sucrose. Comparison of the eight upregulated cDNAs to other sequences in GenBank revealed the following: (1) BSS8A had a 100% identity with the last 25 amino acids of an acyl carrier protein from Arabidopsis thaliana, (2) BSS1A displayed homology to a number of sequences of unknown function, (3) BSS1 IB displayed weak but significant homology to a number of sequences of unknown function, (4) BSS13A displayed homology to four members of the thioredoxin family from ,4. thaliana and (5) four Had no significant homology to previously reported sequences which makes them potential candidates to encode lipogenic enzymes. These results indicate that differential display of mRNA may be a simple and rapid method for the identification of sucrose-modulated gene expression changes in this system and for the characterization of novel sequences potentially encoding lipogenic proteins. / xxi, 256 leaves : ill. ; 28 cm.

No description available.

Previous studies have shown that transcriptional differences in the orf224-atp6 mitochondrial gene region are correlated with fertility restoration of the pol CMS trait by the dominant nuclear Rfp gene in Brassica napus. Recently, the recessive rfp allele, or a tightly linked gene, was found to act as a dominant gene, designated Mmt, in controlling the production of additional, smaller transcripts of two other mitochondrial loci. The results presented in this thesis reveal that Mmt-specific transcripts lack sequences found at the $5 sp prime$ end of the full-length transcripts of these loci and contain a common sequence, UUGUGG, which maps immediately downstream of their $5 sp prime$ termini. A similar sequence, UUGUUG, is found within orf224 downstream of the major Rfp-specific $5 sp prime$ transcript terminus; these hexanucleotide sequences may serve as recognition motifs in the generation of Mmt- and Rfp-specific transcripts. These results suggest that Rfp/Mmt is a novel nuclear locus affecting the expression of multiple mitochondrial gene regions, with different alleles or haplotypes affecting different mitochondrial genes.

Cytoplasmic male sterility (CMS) in plants is associated with mitochondrial dysfunction. We have proposed in this study that the mitochondrial-encoded chimeric peptide thought to be responsible for cytoplasmic male sterility in Polima system could function as a dominant male sterility inducer when expressed in the nucleus and targeted to the mitochondria. Transgenic plants expressing such mitochondrial targeting constructs exhibited reduction of pollen production that was characterized in fertile Westar (nap ) and restored fertile Westar (pol) plants by homeotic transformation of floral organs and in male-sterile Westar (pol) plants by a reduction in pollen production with shortening of the stamens. Genetic and molecular analysis has shown that the phenotypic changes were correlated with the effective genetic transmission of the inserted transgene through female gametes. Most significantly, we have found that differences in floral morphology induced by transgene expression between pol CMS and fertile Westar plants might be related to differences in transcriptional activity of the APETALA3 MADS box gene. We suggest that the alterations in floral morphology that accompany CMS in several plant species might be due to effects of mitochondria on transcriptional activity of floral organ identity genes.

In order to identify organizational differences between sterile Polima (pol) and fertile Campestris (cam) mitochondrial genomes that could be linked to cytoplasmic male sterility (CMS), the physical map of the pol mitochondrial genome was constructed and compared to the physical map of the cam mitochondrial genome. The only structural differences between the two genomes are confined to a region encompassed by a 4.5 kb segment, present in pol mtDNA but absent in cam mtDNA. This 4.5 kb CMS-associated pol segment contains a chimeric gene called orf224 that is cotranscribed with atpG and comprises the single mtDNA region expressed differently in fertile, sterile and fertility restored plants which makes it a good candidate for specifying the sterility trait. Sequence analysis of the pol 4.5 kb segment has shown that orf224 was the only significant open reading frame (ORF) within the segment that gives rise to abundant transcripts, strengthening the view that the orf224/atp6 gene region is conferring pol male sterility. The pol 4.5 kb segment is also present and similarly organized in the common Brassica napus nap mtDNA but the sequences flanking the two segments are unrelated. Thus, the 4.5 kb segment appears to have transposed during the evolution of the pol and nap mitochondrial genomes and appears to have been lost in the cam mitochondrial genome. Sequence analysis of the nap segment revealed the presence of an ORF related to but divergent from orf224. This open reading frame (orf222) potentially encodes a protein of 222 amino-acids with 79% homology to the predicted product of orf224. orf222 is co-transcribed with the third exon of the trans-spliced gene, nad5, and another ORF of unknown function. Expression of the orf222 gene region is tightly associated with nap CMS since the levels of orf222 transcripts are significantly reduced upon restoration while the expression of 22 other mitochondrial genes do not consistently correlate with nap CMS. Antibodies were rai