| Description | The carbon nutritional status in plants and the quality characteristics of agricultural products are often evaluated using sugar content as an important indicator. Monosaccharides and some oligosaccharides (such as maltose) contain free aldehyde or ketone groups, possess reducibility, and are The carbon nutritional status in plants and the quality characteristics of agricultural products are often evaluated using sugar content as an important indicator. Monosaccharides and some oligosaccharides (such as maltose) contain free aldehyde or ketone groups, possess reducibility, and are classified as reducing sugars. Polysaccharides and sucrose are non-reducing sugars. The total sugar content can be determined by measuring the monosaccharide content after hydrolysis, utilizing the property that non-reducing sugars can be hydrolyzed to monosaccharides by acid.Detection Principle: Reducing sugars are oxidized to sugar acids under alkaline heating conditions, while 3,5-dinitrosalicylic acid (DNS) is reduced to a brownish-red amino compound. Within a certain range, the amount of reducing sugar is proportional to the color intensity of the brownish-red product. The absorbance of this brownish-red substance is measured at 540 nm using a microplate reader. This absorbance value has a linear relationship with the reducing sugar content. The reducing sugar and total sugar content in the sample are calculated using a standard curve.This kit is for scientific research use only and is not intended for clinical diagnosis or other purposes.P1501777Component100T300TStorageP1501777AGlu Standard (1 mg/mL)1 mL1 mL2-8℃P1501777BDNS Detection Solution10 mL30 mLRT. Store in the dark.P1501777CColor Solution (for Total Sugar)5 mL10 mLRT. Store in the dark.User-Prepared Instruments and Reagents1. Distilled water, Hydrochloric acid solution, Sodium hydroxide solution2. 50 mL centrifuge tubes, 1 mL centrifuge tubes, Centrifuge, Water bath or incubator, Microplate reader, 96-well plate, Water bathExperimental Procedure1. Extraction of Reducing Sugars1.1 Weigh 0.5-3 g of plant sample, cut into pieces, add about 3 mL of distilled water and homogenize. Transfer to a beaker or conical flask. Rinse the grinder 2-3 times with 12 mL of distilled water and transfer the rinsate to the same container.1.2 Incubate in a 50°C water bath for 30 min, stirring occasionally to ensure thorough extraction of reducing sugars.1.3 Transfer the precipitate and extract to a 50 mL centrifuge tube. Centrifuge at 4000 g for 5 min.1.4 Collect the supernatant. Add 20 mL of distilled water to the precipitate, mix well, and centrifuge again at 4000 g for 5 min.1.5 Collect the supernatant. Combine the supernatants from the two steps. Dilute to 100 mL with distilled water (this is the extract). Mix well. This serves as the test solution for reducing sugars.2. Hydrolysis and Extraction of Total Sugars2.1 Weigh 0.5-3 g of plant sample, cut into pieces, add about 3 mL of distilled water and homogenize. Transfer to a beaker or conical flask. Rinse the grinder 2-3 times with 12 mL of distilled water and transfer the rinsate to the same container.2.2 Add 10 mL of 6 M hydrochloric acid solution to the container, mix well, then heat in a boiling water bath for 30 min for hydrolysis, stirring occasionally.2.3 Take 2 drops and place on a glass slide, add 1 drop of Color Solution (about 50 µL) to check if hydrolysis is complete. If hydrolysis is complete, no blue color should develop.2.4 After hydrolysis, cool to room temperature. Add 6 M sodium hydroxide solution to adjust the pH to 7.4. Dilute to 100 mL with distilled water, mix well. Centrifuge at 4000 g for 5 min or filter.2.5 Take 10 mL of the supernatant or filtrate and dilute to 100 mL with distilled water, creating a 10-fold diluted total sugar hydrolysate (extract). Take 50 µL of this total sugar hydrolysate to measure its reducing sugar content.3. Glucose Standard PreparationTake clean centrifuge tubes or test tubes and prepare a series of Glu standards according to the table below.Standard Working SolutionGlu Standard (1 mg/mL) (mL)Distilled Water (mL)Concentration (mg/mL)10.010.040.220.020.030.430.030.020.640.040.010.850.0501.04. Assay SetupTake 1 mL centrifuge tubes. Set up Blank, Standard, and Test wells according to the table below. Add solutions sequentially, avoiding bubbles. Mix carefully. If the sugar concentration in the sample is too high, reduce the sample volume or dilute appropriately before assay. It is best to set up 2-3 replicate wells for samples and take the average.Reagent (µL)Blank WellStandard WellTest WellDistilled Water50//Glu Standard (1-5)/50/Extract//50DNS Detection Solution100100100Heat accurately in a boiling water bath for 5 min. Remove, cool to room temperature with tap water. Add 250 µL distilled water.5. Reducing Sugar MeasurementMix well. Transfer 300 µL sequentially to the corresponding wells of a 96-well plate. Measure the absorbance of Standard and Test wells at 540 nm, using the Blank well to zero the instrument.6. Result Calculation6.1 Standard Curve PlottingUsing the Glu standards (1-5), i.e., the standard glucose concentrations (mg/mL) as the x-axis and the corresponding absorbance values as the y-axis, plot the standard curve. Find the corresponding glucose concentration on the standard curve based on the absorbance of the extract.6.2 Content CalculationPercentage Content of Reducing Sugars:Reducing sugar content per 100 g sample (g) = (c × V T ) / (m × 1000) × 100 = (c × V T ) / (m × 10)Percentage Content of Total Sugars:Total sugar content per 100 g sample (g) = (c × N × V T ) / (m × 1000) × 100 × 0.9 = (c × N × V T ) / (m × 10) × 0.9Parameter Descriptionc: Sugar amount found from the standard curve (mg/mL)V T : Total volume of the extract, 100 mLm: Mass of the plant sample, gN: Dilution factor of the total sugar hydrolysate, 10Precautions1. Avoid repeated freeze-thaw cycles for the aforementioned low-temperature reagents to prevent inactivation or decreased efficiency.2. If test samples cannot be assayed immediately, store at 2-8°C; stable for 3 days.3. If the sample reducing sugar concentration is too high, dilute with distilled water and re-assay, multiplying the result by the dilution factor.4. The total sugar calculation formula is used when there are few interfering impurities and the reducing sugar content is relatively small compared to the total sugar content. Multiplying by 0.9 accounts for the water consumed during the hydrolysis of total sugars to monosaccharides.5. 6 M Hydrochloric Acid Preparation: Generally, commercially available concentrated hydrochloric acid is 11.6-12 M. Mix concentrated hydrochloric acid with distilled or deionized water 1:1 (v/v) to prepare 6 M HCl. Caution: Hydrochloric acid dissolution in water releases heat; handle carefully to avoid injury.6. 6 M Sodium Hydroxide Preparation: Dissolve 24 g of sodium hydroxide in distilled or deionized water, make up to 100 mL. Caution: Sodium hydroxide dissolution in water releases heat; handle carefully to avoid injury.7. Use reagents promptly after opening to avoid affecting subsequent experimental results... Read More | Product IntroductionAlamar Blue detection reagent provides a simple, rapid, reliable and safe method for cell proliferation and cytotoxicity detection, which is suitable for high-throughput detection experiments. The main component of the detection reagent is a redox indicator. In the oxidized stateProduct IntroductionAlamar Blue detection reagent provides a simple, rapid, reliable and safe method for cell proliferation and cytotoxicity detection, which is suitable for high-throughput detection experiments. The main component of the detection reagent is a redox indicator. In the oxidized state, it appears purple-blue and non-fluorescent, while in the reduced state, it turns into a reduction product with pink or red fluorescence, with an absorption peak of 530-560nm and an emission peak of 590nm.In the process of cell proliferation, the ratios of NADPH/NADP, FADH/FAD, FMNH/FMN and NADH/NAD in the cell increase and are in a reducing environment. The dye taken into the cell is reduced by these metabolic intermediates and cytochromes and then released outside the cell and dissolved in the culture medium, changing the culture medium from non-fluorescent indigo blue to fluorescent pink. Finally, use an ordinary spectrophotometer or fluorophotometer for detection, and the absorbance and fluorescence intensity are proportional to the number of active cells.Instructions1. Add 10µl of detection reagent to 100µl of cell suspension, and incubate in a cell incubator for 2-6 hours. The color of the medium changes from indigo blue to pink and you can proceed to the next step.2. It is recommended to use a fluorescence microplate reader for detection, the excitation light wavelength is between 530-560 nm, the emission light wavelength is 590 nm, and the relative fluorescence unit (RFU) is recorded.3. Draw a standard curve or cell growth curve: the ordinate (Y axis) is the relative fluorescence unit (RFU); the abscissa (X axis) is the cell number or time point or drug concentration.Precautions1. The appropriate density of cells can increase the detection sensitivity. For 96-well plates, we recommend seeding 100 microliters of cells per well. The cell concentration range is: 100-10,000/well for adherent cells, 2,000-50,000/well for suspension cells, and medium as a blank control. For 384-well plates, the cell concentration and seeding volume are both halved.2. The whole process should be aseptic operation, because microbial contaminants can also reduce the detection reagents and affect the experimental results.3. Pay attention to the concentration of inoculated cells and the incubation time after adding detection reagents. If the cell concentration is too high or the incubation time is too long, it will cause a secondary reduction reaction, resulting in colorlessness and disappearance of fluorescence.4. When incubating, avoid light.5. This product can use fluorescence or spectrophotometric detection, but the sensitivity of fluorescence is high, and the experimental error is small. Fluorescence detection is recommended... Read More | Product content C665709Component50 TStorageC665709ABuffer CL45 mLRTC665709BBuffer CB (concentrate)60 mLRTC665709CBuffer GW1 (concentrate)13 mLRTC665709DBuffer GW2 (concentrate)15 mLRTC665709EBuffer EBL10 mLRTC665709FProteinase K100 mgRTC665709GProteinase K Storage Buffer5 mLRTC665709HSpin Columns DFProduct content C665709Component50 TStorageC665709ABuffer CL45 mLRTC665709BBuffer CB (concentrate)60 mLRTC665709CBuffer GW1 (concentrate)13 mLRTC665709DBuffer GW2 (concentrate)15 mLRTC665709EBuffer EBL10 mLRTC665709FProteinase K100 mgRTC665709GProteinase K Storage Buffer5 mLRTC665709HSpin Columns DF with Collection Tubes50 EA2-8℃C665709ICentrifuge Tubes (L-1.5 mL)50 EART Product IntroductionThis kit is suitable for the extraction of free DNA from fresh or frozen serum, plasma, lymph fluid and other cell-free body fluids.This kit adopts centrifugal adsorption columns that can specifically bind nucleic acids and a unique buffer system.After the sample is lysed, the free DNA binds to the silica gel membrane under high salt conditions, and the free DNA elutes from the silica gel membrane at low salt and high pH. The product can handle liquid samples of 0.1-1 ml, and the elution volume of the configured high-efficiency micro adsorption column can be as low as 20 µl. The purified DNA is of high yield and quality, with maximum removal of proteins, pigments, lipids, and other inhibitors, and the rate of free DNA yield is highly dependent on the type of samples, storage conditions, time, and inter-individual variations. The quality of free DNA obtained from purification is stable and reliable, and can be directly used in molecular biology experiments such as PCR, fluorescence quantitative PCR and second generation sequencing.Self-contained reagents: anhydrous ethanol, isopropanol.Pre-experiment Preparation and Important NotesAdd 5 ml of Proteinase K Storage Buffer to Proteinase K to dissolve it and store it at -20℃. Do not leave the prepared Proteinase K at room temperature for a long time.Repeated freezing and thawing of the sample should be avoided, as this can lead to a decrease in extraction.This kit can extract 0.1-1 ml of liquid samples.Before use, please check Buffer CL, Buffer CB for crystallization or precipitation, if there is any crystallization or precipitation, please re-dissolve Buffer CL, Buffer CB by incubation at 56℃ in a water bath.Before first use isopropyl alcohol should be added to Buffer CB according to the instructions on the reagent bottle label, mixed well, and labeled on the reagent bottle label.Before the first use, anhydrous ethanol should be added to Buffer GW1 and Buffer GW2 according to the instructions on the label of the reagent bottle, mixed well, and labeled on the label of the reagent bottle.Preheat the water bath to 60°C before starting the experiment.The elution buffer Buffer EBL can be preheated to 60°C and used.Operation stepsAdd 20 µl of Proteinase K to the centrifuge tube (supplied).Add 200 µl of serum/plasma sample.Note: When the sample volume exceeds 200 µl, please increase the amount of Proteinase K, Buffer CL and Buffer CB reagents in equal proportions, and the specific amount of reagents added can be referred to the attached table.3. Add 160 µl Buffer CL, mix upside down and shake vigorously for at least 30 seconds.4. Incubate at 60°C for 30 minutes, during which time mixing was inverted several times.Note: Incubation of 200µl serum/plasma samples at 60°C for 10-15 minutes is sufficient.Add 360 µl of Buffer CB (check for addition of isopropanol before use) and shake until thoroughly mixed.Ice bath for 5 minutes and centrifuge briefly to concentrate the liquid on the walls and wall caps to the bottom of the tube.Add all of the solution obtained in step 6 to the adsorption columns (Spin Columns DF) that have been loaded into the collection tubes, and if the solution cannot be added all at once, it can be transferred in several times. centrifuge the columns at 12,000 rpm for 1 minute, pour off the waste solution from the collection tubes, and put the columns back into the collection tubes.Add 500µl of Buffer GW1 to the adsorbent column (check that anhydrous ethanol is added before use),centrifuge the column at 12,000rpm for 30 seconds, pour off the waste liquid in the collection tube, and put the adsorbent column back into the collection tube.Add 750 µl Buffer GW2 to the adsorbent column (check that anhydrous ethanol is added before use), centrifuge at 12,000 rpm for 30 seconds, pour off the waste liquid in the collection tube, and put the adsorbent column back into the collection tube.10. Add 750 µl of anhydrous ethanol to the adsorbent column and centrifuge at 12,000 rpm for 30 s. Pour off the waste liquid in the collection tube and put the adsorbent column back into the collection tube.11. Centrifuge at 12,000 rpm for 2 minutes and pour off the waste liquid in the collection tube. Leave the adsorption column at room temperature for several minutes to dry thoroughly.Note: The purpose of this step is to remove residual ethanol from the adsorption column, which can interfere with the subsequent enzymatic reaction.12. Place the adsorption column in a new centrifuge tube, add 20-100 µl Buffer EBL or sterilized water to the middle part of the adsorption column overhanging the column, leave it at room temperature for 2-5 minutes, centrifuge at 12,000 rpm for 1 minute, collect the DNA solution, and store the DNA at -20℃.Note: 1) If the downstream experiment is sensitive to pH, you can use sterilized water for elution. The pH value of the eluent has a great influence on the elution efficiency, if water is used as the eluent should ensure that its pH value is 7.0-8.5 (you can use NaOH to adjust the pH value of water to this range), and the elution efficiency is not high when the pH value is lower than 7.0.2) Preheat the elution buffer BufferEBL to 60℃ and use it, and incubate it at room temperature for 5 minutes before centrifugation to increase the yield.3) If the final concentration of DNA is to be increased, the resulting solution can be reintroduced into the adsorption column and left at room temperature for 2-5 minutes and centrifuged at 12,000 rpm for 1 minute.4) Because DNA preserved in water will be affected by acidic hydrolysis, for long-term storage, it is recommended to elute it with Buffer EBL and store it at -20℃.Table: Recommended reagent additions for different sample sizes... Read More | H665581 Component 100 T Storage H665581A gDNA Eraser 50 µL -20℃. Avoid freeze/thaw cycle. H665581B 10×gDNA Eraser Buffer 120 µL -20℃. Avoid freeze/thaw cycle. H665581C HiFiScript, 200 U/µL 100 µL -20℃. Avoid freeze/thaw cycle. H665581D 5×ScriptRT H665581 Component 100 T Storage H665581A gDNA Eraser 50 µL -20℃. Avoid freeze/thaw cycle. H665581B 10×gDNA Eraser Buffer 120 µL -20℃. Avoid freeze/thaw cycle. H665581C HiFiScript, 200 U/µL 100 µL -20℃. Avoid freeze/thaw cycle. H665581D 5×ScriptRT Buffer 500 µL -20℃. Avoid freeze/thaw cycle. H665581E Primer Mix 120 µL -20℃. Avoid freeze/thaw cycle. H665581F RNase-Free Water 2×1 mL -20℃. Avoid freeze/thaw cycle.Product IntroductionThis product is a kit for removing genomic DNA for reverse transcription. The kit removes genomic DNA in 2 minutes at 42°C. Since the reverse transcription reagent contains a component that inhibits gDNA Eraser, cDNA can be synthesized directly by reverse transcription of gDNA Eraser-treated samples.The kit is equipped with a new high-efficiency reverse transcription enzyme, HiFiScript, with novel mutation sites that dramatically increase the transcriptional activity of the enzyme, resulting in higher efficiency and yield of cDNA first-strand synthesis. The first strand of cDNA can be synthesized with higher efficiency and yield, and the first strand of cDNA can be synthesized from pg total RNA or mRNA. If the reverse transcription product cDNA is used for downstream fluorescence quantitative detection, the reverse transcription reaction can be completed at 42℃ in 15 minutes. This kit is suitable for the synthesis of first-strand cDNA and subsequent RT-PCR, RT-qPCR, and the construction of full-length cDNA libraries.Product Features1. Rapid genome removal: contains gDNA Eraser for genomic DNA removal, which removes genomic DNA in just 2 minutes.2. Rapid reverse transcription: 15 minutes to obtain fluorescent quantitative PCR template cDNA first strand synthesis.3. High sensitivity: cDNA first strand can be synthesized using pg-level total RNA or mRNA templates.4. Highly efficient reverse transcription: Novel mutation sites dramatically increase enzyme activity, resulting in higher yields of cDNA.matters needing attention1. During operation, RNase contamination should be avoided to prevent RNA degradation or cross-contamination in the experiment. It is recommended that operators wear masks and disposable gloves and change the gloves frequently, and use specialized instruments and consumables.2. The reverse transcription system is prepared and operated on ice to prevent degradation of RNA. Store the kit enzymes at -20ºC as soon as possible after use and try to avoid repeated freezing and thawing.3. The reaction system can be scaled up to a maximum of 1 µg of total RNA in 10 µl of reaction system.4. Primer Mix is prepared by Oligo(dT) and Random primer, and Oligo-dT Primer or Gene Specific Primer can also be used according to the experimental needs.5. If the amount of starting RNA is less than 50ng, it is recommended to add RNAase inhibitor (RNasin).6. For RNA templates with complex secondary structures, it is recommended to incubate the template RNA at 65°C for 5 minutes immediately on ice prior to the manipulation step and centrifuge briefly before proceeding to the next step.UsageThaw template RNA on ice; place kit components on ice immediately after thawing at room temperature. Each solution was mixed by vortexing and shaking before use and briefly centrifuged.I. Genomic DNA removal reactions1. Prepare the reaction system according to the following table on ice in a total volume of 10 µl. To ensure the accuracy of the reaction solution preparation, prepare the premixed system in the amount of reaction number + 2 before dispensing it into each reaction tube and finally adding the RNA sample.Note: 1) If the amount of total RNA is greater than 1µg, scale up the reaction system proportionally. If the amount of starting RNA is less than 50ng, it is recommended to add RNAase inhibitor (RNasin).2. Mix by vortex shaking and centrifuge briefly so that the solution on the walls of the tube collects at the bottom.3. Incubate at 42°C for 2 minutes (this can be extended to 30 minutes for room temperature reactions).4．At the end of the reaction, centrifuge briefly and place on ice to cool.II. Reverse transcription reaction1. Prepare the reaction system on ice according to the following table. In order to ensure the accuracy of the reaction solution configuration, first prepare a premixed solution in the amount of number + 2, and then dispense 10 µl into each reaction tube, take 10 µl of the prepared premixed solution and add it to the reaction tube of step 1 where the de-etching of the genome has been completed.Note: 1) Oligo-dT Primer or Gene Specific Primer can be used according to the needs of the experiment, it is recommended to use 50 pmol of Oligo-dT Primer or 2 pmol of Gene Specific Primer for 20 µl reaction system.2. Mix well and centrifuge briefly so that the solution on the walls of the tube collects at the bottom.3. cDNA synthesis reaction conditions:1) If fluorescent quantitative PCR assay is performed downstream, incubate at 42°C for 15 minutes and 85°C for 5 minutes.2) If downstream for normal PCR assay, incubate at 42°C for 30-50 minutes and 85°C for 5 minutes. Note: For templates with complex secondary structure or high GC content, the reverse transcription temperature can be increased to 50°C to enhance reverse transcription efficiency.4. At the end of the reaction, centrifuge briefly and place on ice before proceeding with subsequent PCR or fluorescence quantitative PCR, or place at -20°C if prolonged storage is required.Note: When performing Real-time PCR reactions, the amount of reverse transcription product added should not exceed 1/10 of the total volume of the PCR reaction... Read More | DescriptionTakasago (R)-Ru Cymene Kit I comprises of ruthenium-based biphenyl phosphine cymene catalysts containing either BINAP and SEGPHOS®ligands. These highly reactive and selective catalysts are useful in a variety of asymmetric reactions, mainly asymmetric hydrogenation |