Mitochondrial genetics of alloplasmic male-sterile Brassica napus lines

摘要

Reliable and economical production of hybrid (F1) varieties requires efficient means to control pollination. The predominant system for pollination control of most field crops today is cytoplasmic male sterility (CMS). Since CMS is a consequence of disturbed nuclear-mitochondrial interactions, the CMS trait is maternally inherited. A common way to produce CMS plants is to combine the nuclear genome from one species with the mitochondria from another; plants of such origin are described as alloplasmic CMS. This thesis describes the production and molecular characterization of a novel alloplasmic CMS system produced from rapeseed (Brassica napus) (+) Arabidopsis thaliana somatic hybrids. A population of 170 B. napus (+) A. thaliana somatic hybrid lines was backcrossed to B. napus and 22 lines with male sterility and/or aberrant flower morphology were found. Nine of these were analysed for RFLP and found to contain nuclear and plastid DNA from B. napus, whilst the mitochondrial genomes were frequently recombined containing both B. napus and A. thaliana DNA. Besides the male-sterile trait, the majority of the lines displayed homeotic conversions of anthers to carpeloid organs. Vegetative growth was reduced to some extent in some of the lines. One CMS line segregated both fertile and sterile plants. Fertility co-segregated with molecular markers from A. thaliana chr III. By a dihaploidisation strategy we produced plants with a disomic addition of A. thaliana chr III, which had a stable inheritance of the fertile phenotype. Global mitochondrial mRNA expression was analysed in the fertility-restored line, the corresponding CMS line, B. napus and A. thaliana. Run-on experiments showed that transcriptional activities were highly variable between B. napus and A. thaliana and that in the CMS line transcriptional activity was reduced for several ribosomal protein genes and increased for orf139. Steady-state levels were more homogenous in comparison to transcriptional activities showing that RNA turnover is an important regulatory mechanism. Increased transcript abundance of several genes was observed in the CMS line, often correlated with presence of longer transcripts. Transcripts of three A. thaliana loci, orf139, orf240a and orf294 accumulated in the CMS line, but with reduced levels in the restored line. The orf139 and orf294 transcripts accumulated differentially in a tissue and genotype-specific manner, while orf240a was constitutively expressed throughout the plant. Both orf240a and orf294 transcripts can be polyadenylated, thus providing an explanation for their post-transcriptional regulation. Segregation analysis of sterile and fertile alloplasmic lines indicates that orf139 and orf240a are less likely candidates to be responsible for the male-sterile phenotype, whereas the orf294 can be CMS-associated. However, it is likely that more than one locus in the A. thaliana mt-DNA could encode CMS in the nuclear background of B. napus.