The mid genes of Rhizobium sp strain TAL1145 are required for degradation of mimosine into 3-hydroxy-4-pyridone and are inducible by mimosine.

Mimosine is a toxin present in the tree-legume leucaena (Leucaena leucocephala), including its root nodules and the root exudates. The leucaena-nodulating Rhizobium sp. strain TAL1145 degrades mimosine (Mid(+)) and utilizes it as a source of carbon and nitrogen. Twelve TAL1145 mutants defective in mimosine degradation (Mid(-)) were made through Tn3Hogus, TnphoA or kanamycin-resistance-cassette insertions. A 5.0 kb PstI fragment of TAL1145, subcloned from a cosmid clone containing mid genes for mimosine degradation, complemented most of the Mid(-) mutants. Sequencing this fragment and the adjacent 0.9 kb PstI fragment identified five genes, midA, midB, midC, midD and midR, of which the first three genes encode ABC transporter proteins involved in mimosine uptake, while midD encodes an aminotransferase required for degrading mimosine into 3-hydroxy-4-pyridone, and midR is a regulatory gene encoding a LysR-type transcriptional activator. The location of MidA in the periplasm was shown by making two midA : : phoA fusions, which made active alkaline phosphatase in the periplasm. The various mid : : gus and midA : : phoA fusions were inducible by mimosine, and a midD : : gus fusion mutant showed beta-glucuronidase activity in the leucaena nodules, indicating that midD is expressed in the nodules. Similarly, a midA : : phoA fusion expressed alkaline phosphatase activity in the leucaena nodules, indicating that mimosine induces midA transcription in the bacteroids. mid genes are specific for the Mid(+) strains of leucaena Rhizobium and are absent in strains of other Rhizobium, Sinorhizobium and Bradyrhizobium spp.

Zygotic embryogenesis in Anthurium (Araceae).

Morphological, anatomical, and histochemical aspects of zygotic embryogenesis by Anthurium andraeanum Lind. were investigated from 4 to 24 wk postpollination. Anatomical features were correlated with morphology of the spadix and capacity of embryos to germinate in vitro. Development from a single-cell zygote to fully mature seed takes 24 wk. The suspensor was two ranked and obvious during the early stages of embryogeny. It was apparent by week 8, substantial until week 14, and diminished rapidly until its absence by week 22. Differentiation of the shoot apex, cotyledon, and protoderm occurs at 14 wk. The embryo starts to derive nutrition from the endosperm at this time, and germination of cultured ovules reached 56%. By 20 wk the shoot apex had visible leaf primordia and the root apex was clearly defined. The cotyledon was well developed and surrounded the shoot tip. The storage of protein and starch was at its greatest in the endosperm and embryo. Furthermore, 100% germination of cultured ovules and embryos occurred at 20 wk and thereafter. Fully mature embryos at 24 wk are green and contain protoxylem elements.

Improvements for observing Anthurium (Araceae) floral structures by light microscopy.

Three major obstacles must be overcome in the anatomical study of Anthurium floral structure and embryo development including presence of mucilage, hardened carpel walls, and hardened seed coat in the developing fruit and seed. Fixation in 6% glutaraldehyde and 1% cetylpyridinium chloride in 0.05 M sodium cacodylate, pH 6.8, effectively fixed or removed mucilage from the locules of Anthurium andraeanum Hort. in spadices. This treatment enhanced infiltration of the embedding medium through the hardened carpel wall into the locule space and decreased the quantity and size of holes in the embedding block during sectioning. Specimens 16 weeks after pollination could be fixed, infiltrated, and observed without physical removal of the seed coat. Embryos may be excised from the seed at later stages without compromising embryo structure.

Storage protein changes during zygotic embryogenesis in interior spruce.

The major storage proteins isolated from protein bodies of embryo tissues of interior spruce Picea glauca (Moench) Voss/Picea engelmanii Parry had apparent molecular weights of 41, 35, 33, 24 and 22 kD. Minor proteins of 30 and 27.5 kD were also observed. Based on their solubility characteristics, the 41 kD protein was identified as a water and buffer-soluble albumin, and the 35, 33, 24 and 22 kD proteins were characterized as buffer-insoluble, high salt-soluble globulins. Two-dimensional electrophoresis revealed each protein was composed of several isoelectric variants. Developmentally specific accumulation of storage proteins was observed during embryogenesis. The 41 kD protein only accumulated during the later stages of cotyledon maturation, whereas the other storage proteins began to accumulate during the early stages of embryo development. All storage proteins showed major accumulations during cotyledon maturation.

Characterization of immature embryos of interior spruce by SDS-PAGE and microscopy in relation to their competence for somatic embryogenesis.

SDS-PAGE analysis of total proteins from cotyledonary embryo explants reveals that their competence to form somatic embryos is limited to a specific stage of development prior to the accumulation of storage proteins. When protein profiles of embryo explants of different open pollinated families from the same collection date are compared, there is a close relationship between the absence of storage proteins and their ability to produce embryogenic callus. In addition, the appearance of storage proteins in embryos from subsequent collections is associated with their loss of competence. Light microscopy combined with staining for total protein demonstrates that competent immature embryos have cotyledons but do not contain protein bodies.