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31.
The 45 kDa epidermal growth factor (EGF-(45 kDa)) has been purified from rat urine. We have investigated the distribution and the processing of i.v. injected 125I-labeled EGF-(45 kDa) in the rat. 2.5 min after the i.v. injection only 12% of the label remained in the blood. Most of the label was found in the liver (54%), in the kidneys (7%) and in the skin (4%). The submandibular glands, stomach, small intestine, colon, spleen and lungs contained 1% or less of the radioactivity. Some of the 125I-EGF-(45 kDa) was processed to 125I-EGF-(6 kDa) immunoreactivity in the liver and in the kidneys. The kidneys excreted 125I-EGF-(45 kDa) in the urine, but we were not able to demonstrate 125I-EGF-(6 kDa) in urine. In conclusion, this study shows that homologous EGF-(45 kDa) is cleared from the circulation of rats within a few minutes, mainly by the liver and the kidneys. In vivo both the liver and the kidneys are able to process some of the EGF-(45 kDa) to EGF-(6 kDa) immunoreactivity. 相似文献
32.
Quenching of the fluorescence of DNA-bound Hoechst 33258 in erythroid precursors was studied by flow cytometry and cytochemistry. This quenching artifact may affect the measurement of ploidy in specific cases. The bone marrow cells of two patients with hemolytic disease and active erythropoiesis contained subpopulations of cells with an apparent hypodiploid DNA content as measured by flow cytometry of paraformaldehyde-fixed cells stained with Hoechst 33258. No aneuploidy was detected in either of the two cases when cells were stained with mithramycin or 7-aminoactinomycin D. Cells exhibiting reduced Hoechst 33258 fluorescence expressed glycophorin A and low amounts of CD36, and were therefore erythroid precursors. In one case studied, the number of cells with reduced Hoechst 33258 fluorescence and glycophorin A expressed agreed well with the number of cells containing nuclear hemoglobin. In the other case, hemoglobin was present in a significant proportion of nucleated cells. Calculated values for the efficiency of resonance energy transfer from Hoechst 33258 to hemoglobin were in accordance with the observed levels of quenching (approximately 10%). However, the results could also be explained by hemoglobin reabsorption of Hoechst 33258 fluorescence. Nuclei stained with Hoechst 33258 showed uniform fluorescence, probably due to extraction of hemoglobin during the isolation procedure. 相似文献
33.
Grain yield, symbiotic N2 fixation and interspecific competition for inorganic N in pea-barley intercrops 总被引:4,自引:0,他引:4
Erik Steen Jensen 《Plant and Soil》1996,182(1):25-38
The effect of mixed intercropping of field pea (Pisum sativum L.) and spring barley (Hordeum vulgare L.), compared to monocrop cultivation, on the yield and crop-N dynamics was studied in a 4-yr field experiment using 15N-isotope dilution technique. Crops were grown with or without the supply of 5 g 15N-labeled N m-2. The effect of intercropping on the dry matter and N yields, competition for inorganic N among the intercrop components, symbiotic fixation in pea and N transfer from pea to barley were determined. As an average of four years the grain yields were similar in monocropped pea, monocropped and fertilized barley and the intercrop without N fertilizer supply. Nitrogen fertilization did not influence the intercrop yield, but decreased the proportion of pea in the yield. Relative yield totals (RYT) showed that the environmental sources for plant growth were used from 12 to 31% more efficiently by the intercrop than by the monocrops, and N fertilization decreased RYT-values. Intercrop yields were less stable than monocrop barley yields, but more stable than the yield of monocropped pea. Barley competed strongly for soil and fertilizer N in the intercrop, and was up to 30 times more competitive than pea for inorganic N. Consequently, barley obtained a more than proportionate share of the inorganic N in the intercrop. At maturity the total recovery of fertilizer N was not significantly different between crops, averaging 65% of the supplied N. The fertilizer N recovered in pea constituted only 9% of total fertilizer-N recovery in the intercrop. The amount of symbiotic N2 fixation in the intercrop was less than expected from its composition and the fixation in monocrop. This indicates that the competition from barley had a negative effect on the fixation, perhaps via shading. At maturity, the average amount of N2 fixation was 17.7 g N m-2 in the monocrop and 5.1 g N m-2 in the intercropped pea. A higher proportion of total N in pea was derived from N2 fixation in the intercrop than in the monocrop, on average 82% and 62%, respectively. The 15N enrichment of intercropped barley tended to be slightly lower than of monocropped barley, although not significantly. Consequently, there was no evidence for pea N being transferred to barley. The intercropping advantage in the pea-barley intercrop is mainly due to the complimentary use of soil inorganic and atmospheric N sources by the intercrop components, resulting in reduced competition for inorganic N, rather than a facilitative effect, in which symbiotically fixed N2 is made available to barley.Abbreviations MC
monocrop
- IC
intercrop
- PMC
pea monocrop
- BMC
barley monocrop
- PIC
pea in intercrop
- BIC
barley in intercrop 相似文献
34.
Evaluation of mass spectrometric techniques for charaterization of engineered proteins 总被引:7,自引:0,他引:7
Roepstorff Peter Schram Karl H. Andersen Jens S. Rafn Kate Baldursson Trausti Krøll Jenny Poulsen Kjeld Knudsen Jens Kristiansen Karsten 《Molecular biotechnology》1995,3(1):1-7
A simple and versatile method of in vitro site-specific mutagenesis based on polymerase chain reaction (PCR) is described.
The complete method required the use of three oligonucleotide primers and two PCRs. The product of the first PCR was used
as one of the primers (megaprimer) in the second PCR. Essentially 100% of the final product incorporated the desired mutation.
The various aspects of the procedure and its application is described in detail. 相似文献
35.
Jesper L. Aamand Audrey H. Hobson Catherine M. Buckley Steen T. Jørgensen Borge Diderichsen David J. McConnell 《Molecular & general genetics : MGG》1994,245(5):556-564
An extracellular Pseudomonas cepacia lipase, LipA, is inactive when expressed in the absence of the product of the limA gene. Evidence has been presented that LimA is a molecular chaperone. The lipA and limA genes have been cloned in separate and independently inducible expression systems in Escherichia coli. These systems were used to test the molecular chaperone hypothesis by investigating whether LimA could activate presynthesized prelipase and whether presynthesized LimA could activate newly synthesized prelipase. The results show that LimA cannot activate presynthesized prelipase and that presynthesized LimA can activate only a limited number of de novo synthesized prelipase molecules. Co-immunoprecipitation of prelipase/lipase with LimA generated a 1:1 complex of prelipase/lipase and LimA. The results suggest that a 1:1 complex of LipA and LimA is required for prelipase processing and secretion of active lipase. 相似文献
36.
Charlotte Poulsen Oscar J. M. Goddijn J. Harry C. Hoge Robert Verpoorte 《Transgenic research》1994,3(1):43-49
TransgenicNicotiana tabacum L. Petit Havana SR1 F1-plants expressing tryptophan decarboxylase cDNA (tdc) fromCatharanthus roseus (L.) G. Don under the control of the CaMV 35S promoter and terminator exhibited tryptophan decarboxylase (TDC) enzyme activity and accumulated tryptamine. The plants with the highest TDC activity contained 19 pkat per mg of protein. The influence of transgenic expression oftdc on the activities of anthranilate synthase (AS) and chorismate mutase (CM) were examined in 10 transgenic tobacco plants. The specific activities of these two chorismate-utilizing enzymes were not significantly affected by expression oftdc, despite their important functions as branch point enzymes in the shikimate pathway. The results indicate that the normal route of tryptophan biosynthesis in plants is sufficient to supply a considerable amount of this essential amino acid for the biosynthesis of secondary metabolites. Despite their increased tryptamine content, the growth and development of the transgenic tobacco plants expressingtdc appeared normal. 相似文献
37.
High-affinity folate binding in human prostate 总被引:1,自引:0,他引:1
Binding of3H-folate in Triton X-100 solubilized human prostate homogenate was of a high-affinity type and displayed apparent positive cooperativity typical of specific folate binding. Radioligand dissociation was slow at pH 7.4, but rapid at pH 3.5. Gel chromatography reveled two major folate binding proteins (Mr100 and 25kDa), but only one single band (Mr65–70 kDa) was detectable on SDS-PAGE and immunoblotting with rabbit-anti human milk folate binding protein. Concentration of folate binding protein in prostate homogenate expressed as maximum3H-folate binding was 1.10 nmol/g protein, and the cross-reactivity with rabbit-anti human milk folate binding protein serum was 15% as determined by an enzyme-linked immunosorbent assay (median values; n=6). 相似文献
38.
High-affinity3H-folate binding in Triton X-100 solubilized human mammary gland tissue displayed characteristics, e.g. apparent positive cooperativity and increasing affinity with decreasing concentration of folate binding protein, shown to be typical of specific folate binding. Radioligand dissociation was slow at pH 7.4. A major fraction of the bound radioligand dissociated rapidly at pH 3.5, while a residual binding of 20% persisted even after prolonged dialysis at pH 3.5. Gel chromatography revealed two major folate binding proteins (Mr100 kDa and 25 kDa). However, only one single band was detectable on SDS-PAGE immunoblotting. The highest folate binding activity per g protein was associated with the upper triglyceride-containing layer of the 1000 g supernatant of the homogenate. The folate binding protein extracted from this layer had a low cross-reactivity (<5%) with rabbit antibodies against 25 kDa human milk folate binding protein. The folate binding protein in the 1000 g pellet and the aqueous phase of the 1000 g supernatant was present at a low concentration and had a cross-reactivity of 100%. 相似文献
39.
40.
Jens Chr. Madsen Ole Winneche Sørensen Poul Sørensen Flemming M. Poulsen 《Journal of biomolecular NMR》1993,3(2):239-244
Summary NMR pulse sequences for measuring coupling constants in 13C, 15N-labeled proteins are presented. These pulse sequences represent improvements over earlier experiments with respect to resolution and number of radiofrequency pulses. The experiments are useful for measuring JNH
, JNCO, JNC
, JH
N
CO and JH
N
H
. Applications to chymotrypsin inhibitor 2 (CI-2) are shown. 相似文献