Motility inOscillatoria salina as affected by different factors

All 3-10-d-old Oscillatoria salina filaments glide with the speed of 323-330 microm/min (BG 11 medium, pH 7.5, 21 +/- 2 degrees C, continuous light intensity of approximately 30 micromol m(-2) s(-1)) in a culture chamber. However, a time bound progressive decrease in gliding speed and in percentage of gliding filaments occurred, depending upon the severity of different stress factors studied, viz. water stress (2-8% agarized media, liquid media with 0.2-1 mol/L NaCl, blot-dryness of filaments for > or = 5 min), temperature shock (5, 40 degrees C for > or = 5 min; 35 degrees C for > or = 15 min), darkness and low light intensity (2, 10 micromol m(-2) s(-1)), UV exposure (0.96-3.84 kJ/m2), pH extremes (< or = 6.5 and > or = 9.5), lack of all nutrients from liquid medium (double distilled water), presence of 'heavy' metals (1, 25 ppm Fe, Cu, Zn, Ni, Co, Hg) or organic substances in liquid medium (25, 250 ppm 2,4-D, captan, urea, DDT, thiourea). This feature of the alga (i.e. reduction in speed and percentage of gliding filaments depending upon severity of stress conditions) may thus be suggested to be used in assessing water quality.     

A Thermoelectrically Cooled Microtome Table and Knife

Thermoelectric cooling units (Frigistor thermoelements) have been used to replace CO₂ gas and solid CO₂ for microtome stage and knife cooling. These units consist of assemblies of series-connected thermoelements, functioning by the Peltier effect. Cooling is controlled by the direct current supply to the units. Current supply is from a double power pack giving 15 amp at 4.8 v for the knife cooling units, and 15 amp at 1 v for the stage. By varying the current flow, the optimum cutting temperature can be obtained and held indefinitely. An 8-couple Frigistor unit replaces the CO₂ stage of a Lipshaw freezing microtome. The stage temperature may be lowered to -36 C in 40-60 sec and at the optimum cutting temperature, 5 μ serial sections of fixed frozen tissue are obtainable. Four 12-couple units are used to cool a 160 mm Jung plane wedge microtome knife fitted to a Reichert sledge microtome, with the stage cooled by one 8-couple unit. The knife temperature can be lowered to -20 C in 5 min; the stage in 1 min. The apparatus has been used to cut a variety of unfixed rat and mouse tissues. The optimum sectioning temperature for such unfixed liver, kidney, spleen, lymph glands, heart, testes, small intestine, pancreas, skin and lung was -20 to -22 C.     

Motility and survival ofEuglena ignobilis as affected by different factors

Euglena ignobilis cells in natural puddle water of pH 7.8, when kept at 21 +/- 2 degrees C and under continuous light (intensity of approximately 30 micromol m(-2) s(-1)) in a culture chamber, decreased their speed of movement from > or = 78000 microm/min (after a 12-h cultivation), to 850-1300 microm/min after 18 h. Simultaneously initiated were changes in morphology from the usual elongated motile forms to round motile ones by curving and contraction. Water stress (2 and 4 % agarized puddle water, puddle water with 0.2-1 mol/L NaCl), temperature shock (< or = 10 degrees C, > or = 30 degrees C), darkness and low-light intensity, UV exposure (0.96-2.88 kJ/m2), pH extremes (< or = 6.5 and > or = 10), presence of 'heavy' metals (1-100 ppm Fe, Cu, Zn, Co, Ni, Hg) or organic substances in puddle water (25-1000 ppm 2,4-D, captan, urea, DDT, thiourea), all these factors rapidly (after 5 to 30 min) decreased the speed of the elongated motile form to < or = 300 microm/min, and induced all morphological changes leading to formation of round motile and round nonmotile forms. These features in the alga (i.e. sudden speed reduction and morphological changes from elongate motile to round motile form) may thus be suggested to be used in assessing water quality.     

Thermophilic bacterium Geobacillus uralicus growth is a function of temperature and pH: a synthetic chemostat model-based kinetic analysis

The synthetic chemostat model (SCM), originally developed to describe nonstationary growth under widely varying concentrations of the limiting substrate, was modified to account for the effects of nontrophic factors such as temperature and pH. The bacterium Geobacillus uralicus, isolated from an ultradeep well, was grown at temperatures ranging from 40 to 75 degrees C and at pH varying from 5 to 9. The biomass kinetics was reasonably well described by the SCM, including the phase of growth deceleration observed in the first hours after a change in the cultivation temperature. In an early stage of batch growth in a neutral or alkalescent medium, bacterial cells showed reversible attachment to the glass surface of the fermentation vessel. The temperature dependence of the maximum specific growth rate (micron) was fitted using the equation micron = Aexp(lambda T)/[1 + expB[1-C/(T + 273)]], where A, lambda, B, and C are constants. The maximum specific growth rate of 2.7 h-1 (generation time, 15.4 min) was attained on a complex nutrient medium (peptone and yeast extract) at 66.5 degrees C and pH 7.5. On a synthetic mineral medium with glucose, the specific growth rate declined to 1.2 h-1 and the optimal temperature for growth decreased to 62.3 degrees C.     

Effects of centrifugation, glycerol level, cooling to 5 degrees C, freezing rate and thawing rate on the post-thaw motility of equine sperm

Five experiments evaluated the effects of processing, freezing and thawing techniques on post-thaw motility of equine sperm. Post-thaw motility was similar for sperm frozen using two cooling rates. Inclusion of 4% glycerol extender was superior to 2 or 6%. Thawing in 75 degrees C water for 7 sec was superior to thawing in 37 degrees C water for 30 sec. The best procedure for concentrating sperm, based on sperm motility, was diluting semen to 50 x 10(6) sperm/ml with a citrate-based centrifugation medium at 20 degrees C and centrifuging at 400 x g for 15 min. There was no difference in sperm motility between semen cooled slowly in extender with or without glycerol to 5 degrees C prior to freezing to -120 degrees C and semen cooled continuously from 20 degrees C to -120 degrees C. From these experiments, a new procedure for processing, freezing and thawing semen evolved. The new procedure involved dilution of semen to 50 x 10(6) sperm/ml in centrifugation medium and centrifugation at 400 x g for 15 min, resuspension of sperm in lactose-EDTA-egg yolk extender containing 4% glycerol, packaging in 0.5-ml polyvinyl chloride straws, freezing at 10 degrees C/min from 20 degrees C to -15 degrees C and 25 degrees C/min from -15 degrees C to -120 degrees C, storage at -196 degrees C, and thawing at 75 degrees C for 7 sec. Post-thaw motility of sperm averaged 34% for the new method as compared to 22% for the old method (P<0.01).     

Thermodynamics of carbonic anhydrase catalysis. A comparison between human isoenzymes B and C

The CO2 hydration and HCO3- dehydration activities of human red cell carbonic anhydrase isozymes B and C (HCAB and HCAC) have been studied as a function of temperature from 0 degrees to 37 degrees C. The Arrhenius plots of ln kcat versus 1/T are linear for both isozymes in both hydration and dehydration reactions, indicating that the rate-determining steps remain unchanged over this temperature range. The 37 degrees C hydration kcat, at pH 7.5, is 13 X 10(5) s-1 for isozyme C and 0.71 X 10(5) s-1 for isozyme B. Km, for hydration, is 10 mM for C and 5 mM for B, and invariant with temperature. The uncatalyzed reactions are significantly affected by temperature, 30- to 40-fold rate enhancements being observed from 0 degrees to 37 degrees C. The enzyme-catalyzed processes are much less sensitive to temperature, the rate enhancements being 2- to 3-fold for HCAB and 5- to 6-fold for HCAC in this temperature range. These observations are consistent with a significant lowering of the free energy of activation by both isozymes. This effect is greater for C accounting for its higher catalytic power. The enthalpy of activation, at pH 7.5 and 8.2, in the rate-limiting step is considerably less for the B enzyme compared to C. This is, however, more than offset by a large negative entropy of activation in the case of HCAB. This observation indicates either a mechanistic difference in the rate-limiting events or a difference in the structural organizations of the active sites of the two isozymes, or both.     

Differentiated Staining of Osmium Tetroxide-Fixed, Vestopal-w-Embedded Tissue in thin Sections

Tissues were fixed at 20° C for 1 hr in 1% OsO₄, buffered at pH 7.4 with veronal-acetate (Palade's fixative), soaked 5 min in the same buffer without OsO₄, then dehydrated in buffer-acetone mixtures of 30, 50, 75 and 90% acetone content, and finally in anhydrous acetone. Infiltration was accomplished through Vestopal-W-acetone mixtures of 1:3, 1:1, 3:1 to undiluted Vestopal. After polymerisation at 60° C for 24 hr, 1-2 μ sections were cut, dried on slides without adhesive, and stained by any of the following methods. (1) Mayer's acid hemalum: Flood the slides with the staining solution and allow to stand at 20°C for 2-3 hr while the water of the solution evaporates; wash in distilled water, 2 min; differentiate in 1% HCl; rinse 1-2 sec in 10% NH,OH. (2) Iron-trioxyhematein (of Hansen): Apply the staining solution as in method 1; wash 3-5 min in 5% acetic acid; restain for 1-12 hr by flooding with a mixture consisting of staining solution, 2 parts, and 1 part of a 1:1 mixture of 2% acetic acid and 2% H₂SO₄ (observe under microscope for staining intensity); wash 2 min in distilled water and 1 hr in tap water. (3) Iron-hematoxylin (Heidenhain): Mordant 6 hr in 2.5% iron-alum solution; wash 1 min in distilled water; stain in 1% or 0.5% ripened hematoxylin for 3-12 br; differentiate 8 min in 2.5%, and 15 min in 1% iron-alum solution; wash 1 hr in tap water. (4) Aceto-carmine (Schneider): Stain 12-24 hr; wash 0.5-1.0 min in distilled water. (5) Picrofuchsin: Stain 24-48 hr in 1% acid fuchsin dissolved in saturated aqueous picric acid; differentiate for only 1-2 sec in 96% ethanol. (6) Modified Giemsa: Mix 640 ml of a solution of 9.08 gm KH₂PO₄ in 1000 ml of distilled water and 360 ml of a solution of 11.88 gm Na₂HPO₄-2H₂O in 1000 ml of distilled water. Soak sections in this buffer, 12 hr. Dissolve 1.0 gm of azur I in 125 ml of boiling distilled water; add 0.5 gm of methylene blue; filter and add hot distilled water until a volume of 250 ml is reached (solution “AM”). Dissolve 1.5 gm of eosin, yellowish, in 250 ml of hot distilled water; filter (solution “E”). Mix 1.5 ml of “AM” in 100 ml of buffer with 3 ml of “E” in 100 ml of buffer. Stain 12-24 hr. Differentiate 3 sec in 25 ml methyl benzoate in 75 ml dioxane; 3 sec in 35 ml methyl benzoate in 65 ml acetone; 3 sec in 30 ml acetone in 70 ml methyl benzoate; and 3 sec in 5 ml acetone in 95 ml methyl benzoate. Dehydrated sections may be covered in a neutral synthetic resin (Caedax was used).     

Vitrification of mouse embryos in two cryoprotectant solutions

The objective of this study was to compare the efficiency of 2 media on the vitrification of mouse compacted morulae, early blastocysts and expanded blastocysts after equilibration at room temperature of 4 degrees C. Embryos were equilibrated for 10 min in either 25% VS3 (Rall Equilibration Medium, REM) or 10% glycerol + 20% propylene glycol (Massip Equilibration Medium, MEM) in DPBS at 20 degrees C or 4 degrees C. For vitrification either 100% VS3 (Rall Vitrification Medium, RVM) or 25% glycerol + 25% propylene glycol (Massip Vitrification Medium, MVM) in DPBS was used. Embryos equilibrated at room temperature were loaded in 20 microL of vitrification media into 250 microL straws and then immediately (30 sec) plunged into liquid nitrogen (LN2). After equilibration at 4 degrees C the embryos were put into straws with 20 microL of precooled vitrification medium, and after 20 min at 4 degrees C they were plunged into LN2. Embryos from both groups were thawed in a 20 degrees C water bath for 20 sec, transferred to 1.0 M sucrose in DPBS for 5 min and then cultured for 24 to 48 h in Whitten's medium at 37 degrees C in 5% CO2 in air. In the groups of embryos prepared for vitrification at room temperature the survival rate of compact morulae vitrified in RVM was higher than those vitrified in MVM (65/70, 93% vs 49/74, 66%; P < 0.01). No difference was found in the survival rate of early blastocysts and expanded blastocysts vitrified in RVM or MVM (30/83, 36% vs 25/75, 33% and 4/66, 6% vs 4/76, 5%). No difference was found between the survival rate of compact morulae after equilibration with RVM or MVM at 4 degrees C (62/75, 83% vs 52/74, 70%). Both the early blastocysts and expanded blastocysts equilibrated at 4 degrees C MVM yielded a higher survival rate than RVM (28/74, 38% and 40/70, 57% vs 4/75, 5% and 4/77, 5%; P < 0.01). We conclude that, of the 3 developmental stages, compact morulae withstand the vitrification process best, and reduction of the temperature prior to plunging into LN2 is not required. A 10-fold increase in the survival rate of expanded blastocysts can be achieved using low temperature equilibration (4 degrees C) and MVM.     

产β—葡聚糖酶菌种T199的选育及发酵条件

大麦为啤酒酿造原料 ,含有由葡萄糖残基通过β 1 ,3 和 1 ,4 糖甙键连接而成的β 葡聚糖。在麦芽汁制备过程中 ,热不稳定的大麦葡聚糖酶不能充分降解β 葡聚糖 ,残留的 β 葡聚糖不仅影响麦芽汁分离和啤酒过滤 ,而且将成为成品啤酒出现混浊和沉淀的因素之一。微生物 β 葡聚糖酶能改善啤酒加工工艺和提高产品质量[1,2 ] 。谷类饲料含有不同于纤维素的 β 葡聚糖[2 ] ,作为抗营养因子 ,β 葡聚糖使饲料具有粘性 ,不能很好的消化利用。β 葡聚糖酶作为饲料添加剂加入到饲料中 ,可以将 β 葡聚糖降解 ,从而提高饲料利用率 ,改善营养吸收。相关…     

Staining Paraffin Sections with Protargol 3. The Optimum pH for Reduction. 4. A Two-Hour Staining Method

Further work on conditions affecting the reduction of paraffin sections impregnated with protargol showed that the optimum pH for sulfite-amidol mixtures was between 6.5 and 7.5. A staining method which requires about two hours to complete consists of the following steps: (1) One hour impregnation at 60° C. in 10% AgNO₃. (2) Wash in distilled water 3 changes of 30 sec. each. (3) Put into protargol (Winthrop Chem. Co., New York, N. Y.) 0.2% aq. for another hour at room temperature. (4) Rinse 2 sec. (5) Reduce one to two min. in amidol 0.2 g., Na₂SO₃ 8 g., NaHSO₃ I g., and water 100 cc. (6) Wash thoroly. (7) Tone with 0.1% gold chloride. (8) Wash. (9) Reduce with a 0.5% aq. soln. of amidol (no sulfite). (10) Wash, dehydrate and cover. The method stains neurofibrillae and unmyelinated fibers and has worked well on most tissues of vertebrates. The stain follows acid alcoholic fixation.     

Pre-cooling with intermittent ice ingestion lowers the core temperature in a hot environment as compared with the ingestion of a single bolus

The timing in which ice is ingested may be important for optimizing its success. However, the effects of differences in the timing of ice ingestion has not been studied in resting participants. Therefore, the purpose of this study was to investigate the effects of differences in the timing of ice ingestion on rectal temperature (Tre) and rating of perceptual sensation in a hot environment. Seven males ingested 1.25 g kg⁻¹ of crushed ice (ICE1.25: 0.5 °C) or cold water (CON: 4 °C) every 5 min for 30 min, or were given 7.5 g kgBM⁻¹ of crushed ice (ICE7.5) to consume for 30 min in a hot environment (35 °C, 30% relative humidity). The participants then remained at rest for 1 h. As physiological indices, Tre, body mass and urine specific gravity were measured. Rating of thermal sensation was measured at 5-min intervals throughout the experiment. ICE1.25 continued to decrease Tre until approximately 50 min, and resulted in a greater reduction in Tre (−0.56±0.20 °C) than ICE7.5 (−0.41±0.14 °C). Tre was reduced from 40 to 75 min by ICE1.25, which is a significant reduction in comparison to ICE7.5 (p<.05). Mean RTS with ICE1.25 at 50–65 min was significantly lower than that with ICE7.5 (p<.05). These results suggest that pre-cooling with intermittent ice ingestion is a more effective strategy both for lowering the Tre and for the rating of thermal sensation.     

The effect of pH value and temperature on the stability of L-aminoacidoxidase from the venom of the sand viper]

The stability of highly purified L-amino acid oxidase from the sand viper venom remains practically unaffected by the pH-value at 4degreesC between pH 5 and 8, whereas a sharp activity fall was observed on both sides of this range. At temperatures above 30 degreesC the enzyme is stable only at pH 5.0--5.5. The inactivation pH values above 5.5 follows a first-order rate equation with characteristic changes in the absorption and emission spectra of the enzyme. The stability of the enzyme is dependent on the temperature of storage. At pH 7.5 there is a stability minimum at --10 degrees and -- 30 degreesC. At -- 72 degreesC the enzyme is stable practically for an unlimited period of time; temperatures exceeding 50 degrees C rapidly lead to complete inactivation. Also in the cold, the L-amino acid oxidase is most stable at pH 5.5. There are characteristic changes in absorption and emission spectra in the temperature-stability minimum (--15 degreesC) and at temperatures above 30degreesC. The inactivations follow a first-order rate equation. The cold inactivation is reversible. The stability of the enzyme is diminished by some anions and cations at 37 degreesC. The cold inactivation is promoted by several inorganic anions; organic anions and ammonium sulfate prevent cold inactivation.     

Successful in vitro and in vivo development of in vitro fertilized two- to four-cell cat embryos following cryopreservation, culture and transfer

In vitro and in vivo survival of in vitro-derived 2- to 4-cell cat embryos following cryopreservation was examined. Prefreeze 1- vs 2-step cryoprotectant exposure (Experiment 1) and warming method (Experiment 2) on zona pellucida damage and development in vitro were compared. To determine viability in vivo, frozen/thawed embryos were cultured in vitro to the morula/early blastocyst stage and transferred to synchronous recipients (Experiment 3). At 24 to 26 h after IVF, embryos were cryopreserved in 1.4 M propanediol (Pr) + 0.125 M sucrose (Su) by cooling at 0.3 degrees C/min from -6 degrees C to -30 degrees C and storing in liquid nitrogen. Autologous embryos were cultured in vitro for 7 d. After warming for 5 sec in air and 10 sec at 37 degrees C in water (Experiments 1 to 3), or at room temperature air (22 degrees C; Experiment 2), the cryoprotectant was removed and embryos were cultured in vitro for 6 d (Experiments 1 and 2). Development was assessed after staining by counting cell numbers/embryo and determining the percentages at the 2- to 4-cell (nonsurvivor), pre (5 to 15), early (16 to 32), mid (33 to 50), late (>50) morula or blastocyst stages. Post-thaw development to late morula/blastocyst after 1-step exposure (68%, 15 min Pr + Su) was higher (P< 0.05) than that after 2-step exposure (36%, 15 min Pr and 15 min Pr + Su). Both warming methods produced similar percentages of embryos with damaged zonae (13 to 15%) and equivalent development to morula/blastocyst (64 to 69%). Development in vitro to early morula/blastocyst of frozen embryos with intact zonae was similar to that of nonfrozen embryos. Following cryopreservation, most 2- to 4-cell cat embryos retained their capability for in vitro development to morula/blastocyst, and in vivo viability was demonstrated by the birth of 3 live kittens to 2 of 4 recipients following the transfer of 58 embryos.     

Resistance of pathogenic Naegleria to some common physical and chemical agents.

Resistance of pathogenic Naegleria to drying, low and high temperature, and two halogens was studied. Dying made trophozoites nonviable instantaneously and cysts nonviable in less than 5 min. Trophozoites degenerated in hours at temperatures below 10 degrees C and in minutes when frozen; cysts survived according to the equation th - t0/theta 1,440/1.122T (t0 is survival at 0 degrees C; Tis temperature between 0 and 10 degrees C), but 1.5 h at --10 degrees C to 1 h at --30 degrees C. At 51, 55, 58, 63, and 65 degrees C, trophozoites survived about 30, 10, 5, 1 and less than 0.5 min, respectively, cysts survived three to four times longer at 51 degrees C and six to seven times longer at 55 to 65 degrees C. Cyst destruction rates by heat indicated first-order kinetics with 25,400 cal/1 degree C for energy of activation. Cyst destruction rates by free chlorine and I2 also conformed to first-order kinetics. Concentration-contact time curves yielded concentration coefficient values of 1.05 for free chlorine and 1.4 for I2 and point to superchlorination as an effective means of destroying the cysts if free residuals are used as a guide and allowance is provided for low temperature and/or high pH waters.     

Resistance of pathogenic Naegleria to some common physical and chemical agents

Resistance of pathogenic Naegleria to drying, low and high temperature, and two halogens was studied. Dying made trophozoites nonviable instantaneously and cysts nonviable in less than 5 min. Trophozoites degenerated in hours at temperatures below 10 degrees C and in minutes when frozen; cysts survived according to the equation th - t0/theta 1,440/1.122T (t0 is survival at 0 degrees C; Tis temperature between 0 and 10 degrees C), but 1.5 h at --10 degrees C to 1 h at --30 degrees C. At 51, 55, 58, 63, and 65 degrees C, trophozoites survived about 30, 10, 5, 1 and less than 0.5 min, respectively, cysts survived three to four times longer at 51 degrees C and six to seven times longer at 55 to 65 degrees C. Cyst destruction rates by heat indicated first-order kinetics with 25,400 cal/1 degree C for energy of activation. Cyst destruction rates by free chlorine and I2 also conformed to first-order kinetics. Concentration-contact time curves yielded concentration coefficient values of 1.05 for free chlorine and 1.4 for I2 and point to superchlorination as an effective means of destroying the cysts if free residuals are used as a guide and allowance is provided for low temperature and/or high pH waters.     

Directed evolution of yeast pyruvate decarboxylase 1 for attenuated regulation and increased stability

Five generations of directed evolution resulted in yeast pyruvate decarboxylase 1 (Pdc1) variants with improved activity for 1 mM pyruvate at pH 7.5 in the presence of phosphate. The best variant, named 5LS30, contained the following mutations: A143T, T156A, Q367H, N396I, and K478R. In comparison with native Pdc1, 5LS30 had the substrate concentration required for half-saturation reduced by almost 3-fold at pH 7.5 and the phosphate inhibition reduced by 4-fold at pH 6.0. The apparent cooperativity for pyruvate displayed by 5LS30 was also reduced since it appeared to be activated by pyruvate more easily than the native enzyme. The temperature at which half of the Pdc1 activity was irreversibly lost in 5 min increased from 52.6 degrees C, seen with the native form, to 61.8 degrees C for 5LS30. Curiously, the optimal temperature for Pdc1 activity was found to be dependent upon pyruvate concentration. In 1 mM pyruvate, native Pdc1 performed optimally at 30 degrees C and 5LS30 at 40 degrees C, whereas in 25 mM pyruvate native activity peaked at 45 degrees C and 5LS30 at 55 degrees C. Two screening processes were developed for directed evolution of Pdc1 expressed in Escherichia coli: colony screening and culture screening. The latter proved to be an ideal method for isolating PCR-generated variants of the pdc1 gene with the desired phenotype. In this process, cultures were diluted and partitioned within 96-well plates such that each culture aliquot contained an average of two unique genotypes. This allowed rapid preparation of libraries for analysis of activity in crude lysates and can be applied to other directed evolution projects.     

Zoosporangia survival,dehiscence and zoospore formation,and motility in the green alga<Emphasis Type="Italic">Rhizoclonium hieroglyphicum</Emphasis> as affected by different factors

Urea at 200 ppm (probably serving as a nitrogen source), liquid Bold's basal medium at pH 7.5, temperature of about 22 degrees C and light intensity of about 40 micromol m(-2) s(-1) for 16 h a day induced rapid and/or abundant zoospores formation and zoosporangia dehiscence and favored zoospore liberation, speed and motility time period in the green alga Rhizoclonium hieroglyphicum. However, factors such as water stress (2 and 4 % agarized media, liquid media with 0.2-0.4 mol/L NaCl, 5-60 min blot-dryness of filaments), pH extremes of liquid media (at < or =6.5 and > or =9.5), temperature shock in liquid media (5 and 35 degrees C for > or =5 min), UV exposure (0.96-3.84 kJ/m2), lack of all nutrients from liquid medium (double distilled water), darkness, and presence of "heavy" metals (1-25 ppm Cu, Fe, Zn, Hg, Ni, Co) or organic substances (200-600 ppm captan or DDT, 800 and 1000 ppm 2,4-D, 50 and 400 ppm indole-3-acetic acid (3-IAA), 1000 and 2000 ppm urea, 100 and 200 ppm thiourea) in liquid media decreased and/or delayed at various levels either zoosporangia survival, zoospore formation or zoosporangia dehiscence and/or the rate of zoospore liberation from zoosporangia, zoospore speed and time period of motility in the media or totally inhibited all these processes. 3-IAA at 50 and 400 ppm induced zoosporangial papilla to grow into a tube-like projection of about 30-120 microm in length. Zoosporangial dehiscence rather than zoospore formation or zoosporangia survival, and zoospore motility period rather than zoospore speed are probably more sensitive to various adverse environmental factors. The rate of zoospores liberation from zoosporangium (possibly related directly to some extent on the zoospore number inside) is probably independent of zoospore speed in the medium.     

Rapid Chromosome and Sex-Chromatin Staining with Pinacyanol

This simple and reliable 10-min procedure for producing uniformly and intensely stained, as well as fade-resistant, chromosome and sex-chromatin preparations uses pinacyanol chloride as the dye. Slides are extracted in 5 N HC1 at 20-23 C for 2 min, washed in running tap water for 2 min, stained in 0.25% pinacyanol chloride solution (made up in 70% methanol) for 45 sec, differentiated in Wright's buffer solution (pH 6.4-6.5) for 45 sec, washed in running tap water for 5 sec, dehydrated in 2 changes, 1 min each, of absolute tertiary butanol, cleared in 3 changes of xylene, a minimum of 30 sec each, and mounted in a neutral synthetic resin such as Permount.     

Production and properties of alpha-amylase from Penicillium chrysogenum and its application in starch hydrolysis

Fungi were screened for their ability to produce alpha-amylase by a plate culture method. Penicillium chrysogenum showed high enzymatic activity. Alpha-amylase production by P. chrysogenum cultivated in liquid media containing maltose (2%) reached its maximum at 6-8 days, at 30 degrees C, with a level of 155 U ml(-1). Some general properties of the enzyme were investigated. The optimum reaction pH and temperature were 5.0 and 30-40 degrees C, respectively. The enzyme was stable at a pH range from 5.0-6.0 and at 30 degrees C for 20 min and the enzyme's 92.1% activity's was retained at 40 degrees C for 20 min without substrate. Hydrolysis products of the enzyme were maltose, unidefined oligosaccharides, and a trace amount of glucose. Alpha-amylase of P. chrysogenum hydrolysed starches from different sources. The best hydrolysis was determined (98.69%) in soluble starch for 15 minute at 30 degrees C.     

Enhancement of the Affinity of Nucleoli and Chromosomes for Zinc by Treatment with Chromic Acid

Cultured mammalian cells and wet touch preparations from human organs were fixed for 10 min in 5:85:10 acetic-alcohol-formalin; placed in 5% aqueous CrO₃ for 30 min at 22-25 C; washed in running water 1 min; placed in 2 mM zinc acetate in 0.14 M veronal-acetate buffer, pH 6.5, at 37 C, 30 mm; rinsed 5 sec in 50% acetone; and stained 10 min in a solution dithizone. This results in selective staining of the nucleoli of interphase cells, and of the chromosomes of mitotic cells.