After which the digestions were examined by gel electrophoresis. The samples were run on a 50 mL 0.9% (w/v) agarose gel in 1X TAE buffer at 100 V until the leading track dye traveled 2/3 the distance of the gel. The gel was then soaked in GelRed for 20 minutes and examined under UV light. To prepare the digestions 10 μL of each digestion was mixed with 2 μL of 6X track dye in a micro centrifuge tube. 12 μL of 1 kb DNA ladder and each digestion was run on the
Recovery broth was added to the cell suspension, and the bacteria was placed in warm water for about thirty minutes (see Results and Discussion, paragraph 2). This recovery period let the bacteria repair their cell membranes and express the added genes. Lastly, the transformed E. coli were placed on agar plates and allowed to grow overnight. One agar plate only contained nutrients (-DNA), two contained nutrients and ampicillin, (-DNA/Amp and +DNA/Amp), and one contained nutrients, ampicillin, and IPTG, a protein that caused the GFP to express a glow. After completing the lab, it was discovered that the ARG gene creates a resistance to antibiotics, like ampicillin, and that bacteria can take in new genes.
After the finalization of alkali formulation to be used for extraction of cornhusk fibres, the fibres were treated with Pulpzyme HC to increase their fineness. Pulpzyme HC is a xylanase enzyme (EC.126.96.36.199) produced by submerged fermentation of a genetically modified Bacillus microorganism, and was obtained from Novozymes. The enzyme has an activity of 1000 AXU per gram. (http://fzfz.nbdl.gov.cn:81/files/20130815/1376527490502_36.pdf) Effect of concentration of Pulpzyme HC and Treatment Time The extracted fibres were treated with three different concentrations of enzyme i.e. 1%, 3%, 5% (owf).The time taken for treatment was 30 minutes, 60 minutes and 90 minutes.
MATERIALS AND METHODS Bacterial strains and culture conditions Two S. aureus strains were used in the present study; S. aureus 8325-4 (SigB-) and SH1000 representing a SigB+.strain. Overnight cultures were grown in Luria Broth (LB) at 37°Cwith shaking at 150 rpm. Exposure to antibiotics was carried out as detailed below. Antibiotics Ciprofloxacin were purchased from Sigma-Aldrich CO. 10 mg/ml stock solution of antibiotic were prepared freshly with 0.1N HCl and stored at -20°C. During the experiment we diluted with sterile water 1:10 and 1:100 depending on the different drug concentration.
Potato Dextrose Agar 1. 39.0 g of potato dextrose agar powder was dissolved in a litre of sterile distilled water on the hot plate. 2. pH of the solution was adjusted to 5.6 ± 0.2 by adding NaOH or HCl and was immediately transferred into the Schott bottle to be autoclaved at 121 ° C for 15 minutes 3. Prepared medium was stored in 4° C chiller Plate Count Agar or Total Plate Count 1. 22.5 g of plate count agar powder was dissolved in a litre of sterile distilled water on the hot plate 2. pH of the solution was adjusted to 7.0 ± 0.2 by adding NaOH or HCl and was immediately transferred into the Schott bottle to be autoclaved at 121 ° C for 15 minutes 3.
2.2.Yeast cell preconditioning and inoculum preparation 1 g dry weight of yeast was resuspended in 100 mL of deionized water in an Erlenmeyer flask of 250 mL volume, at 30–35°C, for 30 min with NaCl 6% w/v. Inoculum for experimental fermentations was prepared
Smaller and smaller organelles may be sendimented by successive centrifugation at increasing speed. 1ml of the homogenate (H) was pipetted into a clean microfuge tube labelled P1. The tube labelled ‘P1’ was centrifuged at 1,000g for 10 minutes to sediment the first pellet (P1). The supernatant was carefully removed with an automatic pipette and placed in a micro centrifuge tube labelled ‘P2’. The P2 tube was then placed in the refrigerated centrifuge at a speed of 15,000g for 30-60 minutes at 4°C.
H2SO4 • Catalyst: potassium sulphate (K2SO4), copper (II) sulphate pentahydrate (CuSO4.5H2O) • NaOH (40%) • 0.1 N HCl solution • Boric acid (4%) • Indicator: methyl red (200 mg make up to 100 ml with %95 ethanol ) 1 gram Feta Cheese was used as a sample in this experiment. First of all, the experiment was started with the digestion phase which was performed by mixing 10g potassium sulphate and 0.5 g CuSO4 solution that acts as catalyst, and added 20ml conc. H2SO4. The digestion unit temperature was set to 420±10°C and preheated for 15 minutes. Then, it was heated continuously at least 2 hours till a clear colorless solution was obtained.
21 Fresh semen samples were diluted with phosphate-buffered saline (PBS) to 2x106 sperm/mL. Fifty µL were incubated with 100 µL lysing reagent for 15 seconds and then 2 mL of PI was added and mixed with tube. Tube acquisition was done by flowcytometry immediately after staining. The intensity of its emission corresponds to the DNA content. Flowcytometric analysis displays a constant and characteristic bimodal nonartifactual DNA pattern indicating the presence of two different populations.
A mobile phase system consisting of acetonitrile and 25mM phosphate buffer of pH 3(sodium dihydrogen phosphate monohydrate adjusted with orthophosphoric acid) in a ratio 60:40 (v/v) were used. The mobile phase was degassed and filtered by passing through 0.45 µm pore size membrane filter (Millipore, Milford, MA, USA) prior to use. The flow rate was 1.0 mL min-1 all over the run. The injection volume was 20 µL. The eluent was monitored by the diode array detector (DAD) which was set at 250 nm for the quantitation of both VAL and SAC.
Grow E Coli on an agar plate and grow about 3 of them. Take the e coli colony and swab it with a q tip once swabbed move the e coli to another plate by streaking them across another plate. Incubate the E Coli at 37 degrees celsius for 24 hours. Move the E Coli into the fridge to prevent overgrowth of the E Coli. Place C Elegans into the plates with the E Coli and leave for 24 hours.
In order to do this the scientists will measure the volume of gas that is produced within a 10 second interval time after the tablet begins to react. Then the scientist will observe the different rates of reaction with temperature. The Boltzmann distribution of law, indicates that high temperature makes molecules gain high energy contents (pubs.acs.org/doi/abs/10.1021/ja). In order to measure the reaction rate, the scientists must use the same volume of water at three different starting temperatures: hot tap