Use of labeled nitrogen to monitor transition in nitrogen dependence from storage to current-year uptake in mature walnut trees

Summary Nitrogen (N) derived from both the soil during current-year uptake, and the withintree pool of storage N was distinguished in two groups of Serr walnut trees using labeled fertilizer (¹⁵N-depleted ammonium sulfate) applied in different years. Mass spectrometric analysis of N in xylem sap collected periodically in spring allowed quantification of the relative contributions of N from storage and current season uptake and the transition in N usage from previously assimilated (storage) N to the onset of current season uptake of soil N. N derived from storage accounted for > 50% of the xylem sap N during the staminate and pistillate bloom periods and throughout the period of spur leaf expansion.     

Use of on-line gas analysis to monitor recombinant mammalian cell cultures

The development of a system capable of accurately measuring the oxygen uptake and carbon dioxide production rates during mammalian cell cultures is described. A detailed study of the specifications of the various components used in the system for the measurement of gas flow rates and composition, coupled with the validation of the system independent of the bioreactor was carried out. The aim of this study was to identify and eliminate where possible the errors controlling the accuracy of determination of gaseous metabolic rates. This study showed the importance of controlling the temperature of gaseous oxygen entering the system. With such temperature control, it was possible to obtain data with an accuracy of ±5% at the 95% confidence level. Another source of error, the use of bi-carbonate buffer, was studied. A mathematical model was used to compensate for the affect of such buffers on the determination of catbon dioxide production rates. The use of the system for the continuous determination of gaseous metabolism during the growth and production phase for recombinant CHO cell cultures is described.     

Bioelectrical impedance assay to monitor changes in cell shape during apoptosis

Apoptosis is a strictly regulated and genetically encoded cell 'suicide' that may be triggered by cytokines, depletion of growth factors or certain chemicals. It is morphologically characterized by severe alterations in cell shape like cell shrinkage and disintegration of cell-cell contacts. We applied a non-invasive electrochemical technique referred to as electric cell-substrate impedance sensing (ECIS) in order to monitor the apoptosis-induced changes in cell shape in an integral and quantitative fashion with a time resolution in the order of minutes. In ECIS the cells are grown directly on the surface of small gold-film electrodes (d = 2 mm). From readings of the electrical impedance of the cell-covered electrode, performed with non-invasive, low amplitude sensing voltages, it is possible to deduce alterations in cell-cell and cell-substrate contacts. To improve the sensitivity of this impedance assay we used endothelial cells derived from cerebral micro-vessels as cellular model systems since these are well known to express electrically tight intercellular junctions. Apoptosis was induced by cycloheximide (CHX) and verified by biochemical and cytological assays. The time course of cell shape changes was followed with unprecedented time resolution by impedance readings at 1 kHz and correlated with biochemical parameters. From impedance readings along a broad frequency range of 1-10(6) Hz we could assign the observed impedance changes to alterations on the subcellular level. We observed that disassembly of barrier-forming tight junctions precedes changes in cell-substrate contacts and correlates strongly with the time course of protease activation.     

The use of DNA-intercalating dye to monitor cell fusion and microinjection

A conventional method for microinjection, using erythrocyte ghosts as the injection vector, has been modified to provide a protocol for the highly efficient delivery of small quantities of material into the cytoplasm of target cells. The technique is applicable for use with a variety of proteins, sugars, nucleotides and dyes. When the intercalating dye propidium iodide is included within the sealed ghosts their subsequent fusion with target cells can be continuously monitored by fluorescence spectroscopy, providing a convenient and sensitive parameter of cell-cell fusion. The protocol can be adapted for use with both adherent and non-adherent target cells, and can be used to monitor the relative effectiveness of a variety of fusogenic agents.     

The role of harvest residues to sustain tree growth and soil nitrogen stocks in a tropical Eucalyptus plantation

Aims

Tropical plantations are likely to supply a growing share of the increasing world demand for forest products. We aimed to gain insight into the role of the nitrogen (N) contained in harvest residues (HR) for tree growth and soil N stocks.

Methods

We used 15N-labeled harvest residues to (1) study the dynamic of N release throughout decomposition, (2) determine the vertical transport pathways of N from the forest floor to the upper soil layers, and (3) quantifying the contributions of HR to soil N stocks and the supply of N to young Eucalyptus trees.

Results

Almost all of the 15N initially contained in the HR was recovered 27 months after deposition, with 21 % remaining in HR, 38 % being transferred to the underlying O layer, 21 % being transferred to the 0–15 cm soil layer, and approximately 15 % accumulating in the tree biomass. Our results supported the presence of two pathways of N transfers from the O layer to the mineral soil: (1) the leaching of dissolved 15N from fresh litter during the first year after planting which actively contributed to Eucalyptus N nutrition and (2) the transport of particulate organic matter in percolating water which contributed to maintain N stocks in the first 15 cm of the soil. Approximately 40 % of the N content in 2-year-old Eucalyptus trees was derived from the labeled HR.

Conclusions

The sustainability of fast-growing Eucalyptus trees established on N-poor sandy tropical soils largely relies on organic residues, as an early source of mineral N for tree and as a source of organic N in the top soil.     

Changes in soil organic carbon and total nitrogen stocks after conversion of meadow to cropland in Northeast China

Aims

Grassland conversion to cropland (GCC) may result in loss of a large amount of soil organic carbon (SOC). However, the assessment of such loss of SOC still involves large uncertainty due to shallow sampling depth, soil bulk density estimation and spatial heterogeneity. Our objectives were to quantify changes in SOC, soil total nitrogen (STN) and C:N ratio in 0–100 cm soil profile after GCC and to clarify factors influencing the SOC change.

Methods

A nest-paired sampling design was used in six sites along a temperature gradient in Northeast China.

Results

SOC change after GCC ranged from ?17 to 0 Mg ha^?1 in 0–30 cm soil layer, recommended by IPCC, across the six sites, but ranged from ?30 to 7 Mg ha^?1 when considering 0–100 cm. We found a linear relationship between SOC change in 30–100 cm and that in 0–30 cm profile (ΔC_30?100?=?0.35ΔC_0?30, P?<?0.001), suggesting that SOC change in 0–100 cm was averagely 35 % higher than that in 0–30 cm. The change in STN showed a similar pattern to SOC, and soil C:N ratio did not change at most of sites. On the other hand, SOC loss after GCC was greater in soils with higher initial SOC content or in croplands without applying chemical fertilizers. Furthermore, SOC loss after GCC decreased with falling mean annual temperature (MAT), and even vanished in the coldest sites.

Conclusions

The magnitude of SOC loss following GCC in Northeast China is lower than the global average value, partly due to low MAT here. However, the current low SOC loss can be intensified by remarkable climate warming in this region.     

Survival of shoot meristems of tomato seedlings frozen in liquid nitrogen

An explant containing the primary shoot meristem was dissected from intact tomato seedlings after thawing from liquid nitrogen. Surviving explants produced shoots directly by normal meristem growth when cultured in the presence of gibberellic acid. Without gibberellic acid all surviving explants produced callus tissue and subsequently adventitious shoots, with no direct outgrowth of the primary meristem.Dimethyl sulphoxide (15%) in culture medium and a cooling rate changing continuously from 20 to 55 °C min^?1 between 0 and ?120 °C were required for optimal survival.Nonfrozen material produced shoots directly without the requirement for gibberellic acid indicating that hormonal regulation of organised growth by the shoot meristem had been altered by the freeze/ thaw process.     

Survival of carnation (Dianthus caryophyllus L.) shoot apices frozen to the temperature of liquid nitrogen

Effects of cooling and rewarming rates on the survival of carnationshoot apices frozen to the temperature of liquid nitrogen wereinvestigated. Ten percent dimethyl sulfoxide (DMSO), alone orin combination with 5% glucose, sucrose or sorbitol was mosteffective as a cryoprotectant for carnation shoot apices. Theshoot apices survived slow freezing at about –70?C inthe presence of 10% DMSO. About 80% of the shoot apices survivedfreezing at the temperature of liquid nitrogen after prefreezingat –50?C or below, regardless of the rewarming rates.Shoot apices in the presence of 10% DMSO were cooled at differentrates then rewarmed rapidly. The survival rate gradually decreasedto zero as the cooling rate increased from about 0.5?C/min to50?C/min. At cooling rates higher than 50?C/min, no survivalwas observed even at 5?10⁴?C/min. However, in apices prefrozenat –15?C or below then cooled ultrarapidly at 10⁴?C/min,all remained alive with subsequent rapid rewarming. These apicesdeveloped normal young plants. This ultrarapid cooling methodcombined with prefreezing seems to be useful for the cryopreservationof shoot apices from various plants. ¹Contribution No. 2207 from the Institute of Low TemperatureScience, Hokkaido University. This work was supported in partby a Grant-in-Aid (No. 434035) for Scientific Research fromthe Ministry of Education, Science and Culture. (Received November 13, 1979; )     

Impact of long-term nitrogen addition on carbon stocks in trees and soils in northern Europe

The aim of this study was to quantify the effects of fertiliser N on C stocks in trees (stems, stumps, branches, needles, and coarse roots) and soils (organic layer +0–10 cm mineral soil) by analysing data from 15 long-term (14–30 years) experiments in Picea abies and Pinus sylvestris stands in Sweden and Finland. Low application rates (30–50 kg N ha⁻¹ year⁻¹) were always more efficient per unit of N than high application rates (50–200 kg N ha⁻¹ year⁻¹). Addition of a cumulative amount of N of 600–1800 kg N ha⁻¹ resulted in a mean increase in tree and soil C stock of 25 and 11 kg (C sequestered) kg⁻¹ (N added) (“N-use efficiency”), respectively. The corresponding estimates for NPK addition were 38 and 11 kg (C) kg⁻¹ (N). N-use efficiency for C sequestration in trees strongly depended on soil N status and increased from close to zero at C/N 25 in the humus layer up to 40 kg (C) kg⁻¹ (N) at C/N 35 and decreased again to about 20 kg (C) kg⁻¹ (N) at C/N 50 when N only was added. In contrast, addition of NPK resulted in high (40–50 kg (C) kg⁻¹ (N)) N-use efficiency also at N-rich (C/N 25) sites. The great difference in N-use efficiency between addition of NPK and N at N-rich sites reflects a limitation of P and K for tree growth at these sites. N-use efficiency for soil organic carbon (SOC) sequestration was, on average, 3–4 times lower than for tree C sequestration. However, SOC sequestration was about twice as high at P. abies as at P. sylvestris sites and averaged 13 and 7 kg (C) kg⁻¹ (N), respectively. The strong relation between N-use efficiency and humus C/N ratio was used to evaluate the impact of N deposition on C sequestration. The data imply that the 10 kg N ha⁻¹ year⁻¹ higher deposition in southern Sweden than in northern Sweden for a whole century should have resulted in 2.0 ± 1.0 (95% confidence interval) kg m⁻² more tree C and 1.3 ± 0.5 kg m⁻² more SOC at P. abies sites in the south than in the north for a 100-year period. These estimates are consistent with differences between south and north in tree C and SOC found by other studies, and 70–80% of the difference in SOC can be explained by different N deposition.     

Survey of ATCC stocks of human cell lines for HeLa contamination

Seed stocks of human cell lines deposited at the American Type Culture Collection (ATCC) have been examined for cross-contamination with HeLa cells using Giemsabanded marker chromosomes. Sixteen additional cell lines investigated have been found to exhibit marker chromosomes typical of HeLa cells. Quinacrine fluorescence studies further revealed the absence of Y chromosome in these lines. These observations indicate that the lines are HeLa derivatives.     

Survey of ATCC stocks of human cell lines for hela contamination

Summary Seed stocks of human cell lines deposited at the American Type Culture Collection (ATCC) have been examined for cross-contamination with HeLa cells using Giemsabanded marker chromosomes. Sixteen additional cell lines investigated have been found to exhibit marker chromosomes typical of HeLa cells. Quinacrine fluorescence studies further revealed the absence of Y chromosome in these lines. These observations indicate that the lines are HeLa derivatives.