Expression of the [beta]-Glucuronidase Gene in Pollen of Lily (Lilium longiflorum), Tobacco (Nicotiana tabacum), Nicotiana rustica,and Peony (Paeonia lactiflora) by Particle Bombardment

A [beta]-glucuronidase (GUS) gene that is under the control of the anther-specific LAT52 promoter of tomato (Lycopersicon esculentum) and the nopaline synthetase polyadenylation terminator was successfully expressed in pollen of Lilium longiflorum, Nicotiana tabacum, Nicotiana rustica, and Paeonia lactiflora using a pneumatic particle gun. The GUS gene in plasmid pBI221 was also expressed, to a lesser extent, in pollen of all of these species. The presence of methanol in the substrate solution for histochemical GUS assay and the incubation time in this solution influenced successful detection of GUS expression in bombarded pollen. Cytological analysis of GUS-expressing pollen of lily showed that introduced gold particles were seen in intracellular compartments of pollen, including the vegetative cytoplasm, vegetative nucleus, and generative cytoplasm.     

On the Function of Mitochondrial Metabolism during Photosynthesis in Spinach (Spinacia oleracea L.) Leaves (Partitioning between Respiration and Export of Redox Equivalents and Precursors for Nitrate Assimilation Products)

The functioning of isolated spinach (Spinacia oleracea L.) leaf mitochondria has been studied in the presence of metabolite concentrations similar to those that occur in the cytosol in vivo. From measurements of the concentration dependence of the oxidation of the main substrates, glycine and malate, we have concluded that the state 3 oxidation rate of these substrates in vivo is less than half of the maximal rates due to substrate limitation. Analogously, we conclude that under steady-state conditions of photosynthesis, the oxidation of cytosolic NADH by the mitochondria does not contribute to mitochondrial respiration. Measurements of mitochondrial respiration with glycine and malate as substrates and in the presence of a defined malate:oxaloacetate ratio indicated that about 25% of the NADH formed in vivo during the oxidation of these metabolites inside the mitochondria is oxidized by a malate-oxaloacetate shuttle to serve extramitochondrial processes, e.g. reduction of nitrate in the cytosol or of hydroxypyruvate in the peroxisomes. The analysis of the products of the oxidation of malate indicates that in the steady state of photosynthesis the activity of the tricarboxylic acid cycle is very low. Therefore, we have concluded that the mitochondrial oxidation of malate in illuminated leaves produces mainly citrate, which is converted via cytosolic aconitase and NADP-isocitrate dehydrogenase to yield 2-oxoglutarate as the precursor for the formation of glutamate and glutamine, which are the main products of photosynthetic nitrate assimilation.     

Current Nitrogen Fixation Is Involved in the Regulation of Nitrogenase Activity in White Clover (Trifolium repens L.)

Previous studies have shown that nitrogenase activity decreases dramatically after defoliation, presumably because of an increase in the O2 diffusion resistance in the infected nodules. It is not known how this O2 diffusion resistance is regulated. The aim of this study was to test the hypothesis that current N2 fixation (ongoing flux of N2 through nitrogenase) is involved in the regulation of nitrogenase activity in white clover (Trifolium repens L. cv Ladino) nodules. We compared the nitrogenase activity of plants that were prevented from fixing N2 (by continuous exposure of their nodulated root system to an Ar:O2 [80:20] atmosphere) with that of plants allowed to fix N2 (those exposed to N2:O2, 80:20). Nitrogenase activity was determined as the amount of H2 evolved under Ar:O2. An open flow system was used. In experiment I, 6 h after complete defoliation and the continuous prevention of N2 fixation, nitrogenase activity was higher by a factor of 2 compared with that in plants allowed to fix N2 after leaf removal. This higher nitrogenase activity was associated with a lower O2 limitation (measured as the partial pressure of O2 required for highest nitrogenase activity). In experiment II, the nitrogenase activity of plants prevented from fixing N2 for 2 h before leaf removal showed no response to defoliation. The extent to which nitrogenase activity responded to defoliation was different in plants allowed to fix N2 and those that were prevented from doing so in both experiments. This leads to the conclusion that current N2 fixation is directly involved in the regulation of nitrogenase activity. It is suggested that an N feedback mechanism triggers such a response as a result of the loss of the plant's N sink strength after defoliation. This concept offers an alternative to other hypotheses (e.g. interruption of current photosynthesis, carbohydrate deprivation) that have been proposed to explain the immediate decrease in nitrogenase activity after defoliation.     

Identification and Characterization of a Biodegradative Form of Threonine Dehydratase in Senescing Tomato (Lycopersicon esculentum) Leaf

Threonine dehydratase (TD; EC.4.2.1.16) is a key enzyme involved in the biosynthesis of isoleucine. Inhibition of TD by isoleucine regulates the flow of carbon to isoleucine. We have identified two different forms of TD in tomato (Lycopersicon esculentum) leaves. One form, present predominantly in younger leaves, is inhibited by isoleucine. The other form of TD, present primarily in older leaves, is insensitive to inhibition by isoleucine. Expression of the latter enzyme increases as the leaf ages and the highest enzyme activity is present in the old, chlorotic leaves. The specific activity of the enzyme present in older leaves is much higher than the one present in younger leaves. Both forms can use threonine and serine as substrates. Whereas TD from the older leaves had the same Km (0.25 mM) for both substrates, the enzyme from the young leaves preferred threonine (Km = 0.25 mM) over serine (Km = 1.7 mM). The molecular masses of TD from the young and the old leaves were 370,000 and 200,000 D, respectively. High levels of the isoleucine-insensitive form of threonine dehydratase in the older leaves suggests an important role of threonine dehydratase in nitrogen remobilization in senescing leaves.     

Are viruses important partners in pelagic fend webs?

Viruses have been assumed to play a rather negligible role as partners in microbial food web dynamics. However, recent discoveries suggest that the rate of virally induced lysis of marine microbial populations may be significant. This, in turn, may have important consequences for the developing conceptual framework of the microbial food web.     

Effect of ovarian antral follicle cauterization on the interestrus interval of the gilt

The objective of the present study was to determine if destruction of ovarian antral follicles by laser-cauterization affects CL lifespan during the estrous cycle of the gilt. Cyclic gilts were randomly assigned to either SHAM, laser (L) or laser-estradiol (L-E2) treatment groups, with the L-E2 group receiving a 5-mg intramuscular (i.m.) injection of estradiol-17beta cypionate at the time of the first surgery. Ovarian antral follicles were laser-cauterized on either Days 12 and 14 (L12) or Days 14 and 17 (L14) of the estrous cycle. In the L12-E2 group, 3 of 4 gilts had extended mean interestrus intervals of more than 22 days compared with 0 of 4, 0 of 6, 0 of 7 and 1 of 5 gilts in the SHAM, L12, L14 and L14-E2 groups, respectively. The L12-E2 gilts had a longer (P<0.05) mean interestrus interval (23.5+/-1.3 days) than the L12 (20.0+/-1.1 days), L14 (20.7+/-1.0 days) and SHAM (20.5+/-1.3 days). The mean interestrus interval of L14-E2 gilts (21.8+/-1.2 days) did not differ from those of the L12-E2 group or the L12, L14 and SHAM group gilts. Six additional gilts were injected with 5 mg estradiol cypionate-17beta to serve as nonsurgical controls for E2 treatment. Gilts (3 of 3) given an E2 injection on Day 12 had extended mean interestrus interval (26.0+/-2.6 days), while 2 of 3 gilts injected with E2 on day 14 had extended mean interestrus intervals (27.7+/-2.1 days). These results indicate that in cyclic gilts destruction of ovarian follicles by laser-cauterization did not affect CL lifespan, and that luteolysis is not dependent on the presence of antral follicles.     

In vitro survival of murine morulae after quick freezing in the presence of chemically defined macromolecules and different cryoprotectants

We studied the ability of frozen-thawed mouse morulae to develop in vitro when the cryoprotectant proteins were substituted with one of the following nonorganic macromolecules: polyvinyl alcohol (PVA), polyvinyl pyrrolidone (PVP), and ficoll. We also determined how these agents interacted with 3 different cryoprotectants: glycerol (GLY), propylene glycol (PG), and ethylene glycol (EG). The influence of both of the above factors was measured on the basis of post-thaw morphological appearance, the percentage of development to the expanded blastocyst stage and the total cell count. Morulae (n=950) were collected from superovulated mice. Those classified as good or excellent were distributed among the 12 different freezing solutions, obtained by combining the 3 cryoprotectants with the 4 macromolecules (the 3 mentioned above, plus a control of 5% fetal calf serum) in phosphate buffered saline (PBS). Embryos frozen in PVA, PVP and ficoll tended to be a little difficult to recover from the straws. Development to the expanded blastocyst stage was significantly lower (P<0.05) in propylene glycol (43.6%) than in ethylene glycol (79.5%) or in glycerol (76.1%). Polyvinyl alcohol provided a higher survival rate when combined with glycerol (90.3) or ethylene glycol (95.0), but when it was combined with propylene glycol, only 56.5% of embryos survived after thawing. A positive interaction was observed between glycerol and PVA and between ethylene glycol and PVA or ficoll. The results indicate that fetal serum could be successfully substituted for any of the 3 chemically defined macromolecules. However, our findings also suggest that the use of PG as a cryoprotectant should be avoided when mouse morulae are frozen using the quick freezing method.     

毛喉萜对小鼠骨髓细胞和腹腔巨噬细胞表面胰岛素受体和GM－CSF受体的下调作用

放射性标记受体分析表明毛喉萜（ＦＳＫ）可以降低小鼠骨央细胞和腹腔巨噬细胞表面的胰岛素（ＩＮＳ）和粒细胞－巨噬细胞集落刺激因子（ＧＭ－ＣＳＦ）受体数目,而且对ＧＭ－ＣＳＦ的作用有剂量和时间依赖性,经ＦＳＫ处理的巨噬细胞酸性磷酸酶活性增加,微丝更加舒展。     

赤松林受虫害后生物学及光谱学特征的变化

 本文重点研究春季山东招远赤松林受赤松毛虫危害后,其针叶形态特征、叶绿素含量及光谱反射率特征的变化,结果表明：春季赤松受赤松毛虫啃食危害后,树势衰弱,生长不良,针叶颜色变化,褪绿甚至死亡,可分为五个等级;随受害程度的加重,叶绿素含量逐渐降低;赤松受损后,红界光谱曲线拐点“蓝移”,可用特征参量Rs、Ro、λp、λpr、λo、R550-680的值表征。