Complex organization of the mitochondrial genome of petaloid CMS carrot.

The petaloid trait in carrot is a mitochondrially associated homeotic-like conversion of stamens into petals which results in cytoplasmic male sterility (cms), but little is understood about the phenomenon at the molecular level. Identification of region(s) of the mitochndrial (mt) genome that are causally implicated in cms may be aided by a physical map of cms-associated mtDNA. The mt genome of petaloid cms carrot is 255 kb in length and contains three pairs of repeated sequences, two in direct orientation and one in inverted orientation. All regions of the genome are present in equal stoichiometries, but the arrangement of the repeated sequences prevented the representation of the entire genome as a single master circle or multiple subgenomes. An alternative model of mtDNA replication that accounts for our data is discussed.

Interactions of ancymidol with sucrose and α-naphthaleneacetic acid in promoting asparagus (Asparagus officinalis L.) somatic embryogenesis.

Interactions of varying ancymidol concentrations with those of α-naphthaleneacetic acid (NAA) or sucrose in embryo induction medium were related to the production and development of asparagus (Asparagus officinalis L.) somatic embryos, and to the ability of these embryos to germinate. A significant sucrose×ancymidol interaction was observed only for the production of bipolar embryos; 4% sucrose with 0.75 mg l-1 ancymidol gave the best result, 78 g-1 callus. The frequency of globular embryos decreased as sucrose or ancymidol concentrations increased. Sucrose concentration affected embryo germination; 3% and 4% sucrose were optimal with approximately 60% and 40% of bipolar and globular embryos germinating, respectively. Significant ancymidol×NAA interactions were observed for the production of bipolar and globular embryos and their germination. Varying ancymidol concentrations affected embryo production and germination in combination with 0.1 mg l-1 NAA, but not with 1.0 mg l-1 NAA. The treatment combination of 0.1 mg l-1 NAA with 0.75 mg l-1 ancymidol produced the most bipolar embryos, 64 g-1 callus, and the greatest percentages of bipolar and globular embryos germinated, 63% and 42%, respectively.

Chromosome doubling via tuber disc culture in dihaploid potato as determined by confocal microscopy.

The potential of tuber disc culture for chromosome doubling was investigated in somaclonal populations of four dihaploid genotypes and one tetraploid cultivar of potato (Solanum tuberosum). Laser scanning confocal microscopy (LSCM) was used for rapid determination of the ploidy level based on the number of chloroplasts in stomatal guard cells of leaves. Factorial analysis of chloroplast number in 58 clones and two leaf types showed that somaclones were clearly divided in two groups. Clones with 5-7 chloroplasts per cell as observed in tuber derived diploid controls were classified as 2X (not doubled), while those with 9-14 chloroplasts resembled the tuber derived tetraploid controls and were considered 4X (doubled). A high frequency of spontaneous chromosome doubling, 42% - 50%, was detected in 3 dihaploid genotypes, whereas no doubling was observed in one of the dihaploids as well as the tetraploid cultivar Yukon Gold. Effects of leaf type on chloroplast number was also significant. The middle leaf showed significantly higher chloroplast number than the younger leaves.

The effects of ancymidol, abscisic acid, uniconazole and paclobutrazol on somatic embryogenesis of asparagus.

The effects of ancymidol, abscisic acid (ABA), uniconazole, and paclobutrazol on asparagus somatic embryogenesis were evaluated. Calli induced from seedlings of genotype G447 were transferred to embryo induction medium (MS plus 3% sucrose, 0.1 mg L(-1) NAA, 0.5 mg L(-1) kinetin and 3% gelrite), with different concentrations of these compounds. After 8 weeks, the recovered bipolar or globular embryos were placed on germination medium (MS plus 6% sucrose, 0.1 mg L(-1) NAA, 0.1 mg L(-1) kinetin, 0.75 mg L(-1) ancymidol, 40 mg L(-1) adenine sulphate dihydrate, 0.17 mg L(-1) sodium phosphate monobasic and 3% gelrite) for conversion to plantlets. Inclusion of ancymidol, ABA, uniconazole and paclobutrazol in the embryo induction medium did not affect the total number of somatic embryos produced relative to the control without these compounds. However, ancymidol, ABA and uniconazole significantly improved embryo development by increasing the production of bipolar embryos 250-750% and decreasing that of globular embryos 8-35% relative to the control. The bipolar embryos produced with any of the four compounds in the embryo induction medium converted to plantlets at rates 700-1100% greater than the control. None of the globular embryos converted to plantlets. Ancymidol (0.75 mg L(-1)) and ABA (0.05 mg L(-1)) were the most effective treatments; 61 and 46 bipolar embryos g(-1) callus were produced, and 38% and 37% of the bipolar embryos converted to plantlets, respectively. These results indicated that ancymidol, ABA, uniconazole and paclobutrazol significantly enhanced the production of asparagus somatic embryos and their conversion to plantlets, and ancymidol and ABA were more effective than uniconazole and paclobutrazol.

Development of haploid asparagus embryos from liquid cultures of anther-derived calli is enhanced by ancymidol.

The effect of ancymidol concentration on the development of haploid asparagus embryos was determined. Liquid cultures from anther-derived calli were grown for three weeks in MS medium plus 1.0 mg l(-1) 2,4-D, 0.1 mg l(-1) NAA, 0.2 mg l(-1) kinetin, 800 mg l(-1) glutamine, 500 mg l(-1) casein hydrolysate, 2% sucrose and 0.0-1.0 mg l(-1) ancymidol. Cell clumps (224-500 µm) were plated on solid embryo maturation medium (MS medium plus 3% sucrose, 0.1 mg l(-1) NAA, 0.5 mg l(-1) kinetin and 0.0-1.0 mg l(-1) ancymidol) and grown for eight weeks. Ancymidol enhanced embryo maturation and germination and was more critical in the solid than liquid medium. Total embryo number did not vary among most treatments. The best response was observed when ancymidol concentrations were 0.1 and 0.5 mg l(-1) in the liquid and solid media, respectively; two-thirds of the embryos produced were bipolar and 35% of bipolar embryos germinated. Seven to 82% of plants recovered from different ancymidol treatments were haploid; the others were diploid, triploid or chimeric for ploidy level.

High frequency production of haploid embryos in asparagus anther culture.

A method for obtaining a high frequency of haploid asparagus embryos through anther culture was developed. Flowers collected from plants in the field in July, August and September 1990, for the genotype G203, were stored at 5°C for 24 h. Anthers were placed on Murashige and Skoog medium (MS) containing 500 mg l (-1) casein hydrolysate, 800 mg l(-1) glutamine, 2 mg l (-1) NAA, 1 mg l (-1) BA and 5 % sucrose at 32 °C in the dark for three to four weeks to induce calli. Calli were then grown at 25 °C with a 16 h photoperiod for three to four weeks. Developing embryos and calli were transferred to embryo maturation medium, MS containing 6% sucrose, 0.1 mg l (-1) NAA, 0.1 mg l (-1) kinetin and 0.65 mg l (-1) ancymidol, for four weeks. More than 50% of the recovered mature embryos germinated on MS containing l mg l (-1) GA3. Anthers with microspores at the late-uninucleate stage had the highest frequency of total and embryogenic calli formation, 40% and 15%, respectively. Each embryogenic callus usually produced 10-15 embryos. Aproximately 75 plants per 100 anthers cultured were recovered: 76% haploid, 22% diploid and 2% triploid. High temperature was critical for the induction of embryogenic callus.