S1 and S2 are short linear mitochondrial DNA molecules found in a particular male sterile cytoplasm of maize. We show here that these DNA molecules and two other related linear DNA species found in maize mitochondria, have proteins attached, probably covalently, to their 5' ends. This is the first demonstration of such a linear DNA-terminal protein association in higher eukaryotes. Such proteins may be involved in priming replication of these DNAs.
In microspore cultures of Brassica napus L. cv. Topas, embryo yield increases with culture density up to about 40,000 microspores per ml. A much higher density (100,000 per ml) appears inhibitory to embryogenesis. A relatively high culture density (30,000 or 40,000 per ml) for the first 2-4 days of culture is crucial for embryogenesis, after which cultures may be diluted to allow better embryo growth.Medium conditioned by culturing microspores at 30,000 or 40,000 per ml for 1 day improved microspore-embryo yield in low density cultures (3,000 or 4,000 per ml) more than 3-fold. In contrast, media conditioned with microspores from 1-4 days or 0-4 days of culture were inhibitory.Use of feeder cultures resulted in up to 10-fold increase of embryo yield in low density microspore cultures, depending on the method used. Filter papers and other membranes placed on top of feeders greatly inhibited embryogenesis in the feeder layer as well as microspores cultured on the feeder, possibly due to poorer gaseous exchange.
The durations of meiosis and pollen development were estimated in Petkus Spring rye and Chinese Spring wheat grown at 15, 20 and 25 °C. Over the range 15-25 °C, meiotic duration and pollen maturation time decreased with increasing temperature and the ratio between the durations of these stages at 25 °C and at 15 °C (Q10) indicated that the temperature responses of meiotic and pollen development were similar in both species. Both within and between species meiosis and pollen maturation had constant relative durations at each temperature. The effect of low temperature on the duration of meiosis and pollen development was similar in Holdfast, an extreme type of winter wheat, as in Chinese Spring. The relevance of these results to hybrid cereal breeding are mentioned. In plants grown at 25 °C, abnormal meiotic and pollen development resulting in male sterility was seen in some anthers. The cause of male sterility appeared to be thickening of tapetal cell walls concurrent with abnormal late meiotic development in PMC's.
An assay is described whereby Eco RI restriction fragment length polymorphisms of mitochondrial and chloroplast DNAs can definitively identify cytoplasms of interest in Brassica crop development. Restrictable mitochondrial and chloroplast DNA is extracted from as little as 2-3 g and 0.5 g leaf tissue, respectively, and the donor plants are able to continue to develop in a normal manner. An unknown cytoplasm can be identified in three days, which is a considerable saving in time and labor compared to the several years required by traditional methods. The assay is very inexpensive and should be established as a routine procedure in laboratories involved in sexual or somatic Brassica hybrid production.
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