ABSTRACT
A cytoplasmic male sterile (CMS) line of Brassica juncea was derived by repeated backcrossing of the somatic hybrid (Diplotaxis catholica + B. juncea) to B. juncea. The new CMS line is comparable to euplasmic lines for almost all characters, except for flowers which bear slender, needle-like anthers with aborted pollen. Detailed Southern analysis revealed two copies of coxI gene in the CMS line. One copy, coxI-1 is similar to the coxI gene of B. juncea, whereas the second copy, coxI-2 is present in a novel rearranged region. Northern analysis with eight mitochondrial gene probes showed altered transcript pattern only for the coxI gene. Two transcripts of 2.0 and 2.4 kb, respectively, were detected in the CMS line. The novel 2.4 kb transcript was present in floral bud tissue but absent in the leaf tissue. In plants where male sterility broke down under high temperature during the later part of the growing season, the 2.4 kb coxI transcript was absent, which suggested its association with the CMS. The two coxI genes from the CMS line showed two amino acid changes in the coding region. The novel coxI gene showed unique repeats in the 5' region suggesting recombination of mitochondrial genomes of the two species. The possible role of the duplicated coxI gene in causing male sterility is discussed.
Subject(s)
Base Sequence , Brassica/genetics , Cyclooxygenase 1/genetics , Cytoplasm/genetics , DNA, Mitochondrial/analysis , Flowers/genetics , Gene Duplication , Gene Expression , Genome, Plant , Hybrid Cells/metabolism , Molecular Sequence Data , Mustard Plant/genetics , Plant Infertility/genetics , RNA/analysis , Random Amplified Polymorphic DNA Technique , Sequence Homology, Nucleic AcidABSTRACT
A number of factors that are known to influence genetic transformation were evaluated to optimize Agrobacterium-mediated transformation of hypocotyl explants of cauliflower variety Pusa Snowball K-1. The binary vector p35SGUSINT mobilized into Agrobacterium strain GV2260 was used for transformation and transient GUS expression was used as the basis for identifying the most appropriate conditions for transformation. Explant age, preculture period, bacterial strain and density were found to be critical determinants of transformation efficiency. Using the optimized protocol, the synthetic cryIA(b) gene was mobilized into cauliflower. Molecular analyses of transgenics established the integration and expression of the transgene. Insect bioassays indicated the effectiveness of the transgene against infestation by diamondback moth (Plutella xylostella) larvae
Subject(s)
Brassica/genetics , Plants, Genetically Modified , Rhizobium/genetics , Transformation, GeneticABSTRACT
Plants were regenerated from callus induced from leaf disc explants of a tomato F1 hybrid heterozygous for three marker loci anthocyaninless (a), without anthocyanin (aw), and hairless (hl). Regenerants were studied for somaclonal variation at the phenotypic level by scoring for variation in the marker loci, and at the DNA level by probing geomic DNA blots with a chlorophyll a/b binding protein (Cab-3C) cDNA sequence. While no variation was observed at the phenotypic level in over 950 somaclones studied, DNA polymorphism for the Cab locus could be detected in two out of 17 somaclones tested. Tissue culture induced variation at the phenotypic level for specific loci is very low (less than 0.001 for a, aw or hl) but DNA sequence changes are induced at much greater frequency (approximately 0.1 for a multicopy gene family such as Cab).