| Description | DescriptionIt contains a set of seven different homogeneous palladium catalysts, useful for rapid screening of catalysis conditions. It is in sampler format with individual components packaged for multiple experiments and mini scale-up. The cost of the kit is less than the total cost of individual DescriptionIt contains a set of seven different homogeneous palladium catalysts, useful for rapid screening of catalysis conditions. It is in sampler format with individual components packaged for multiple experiments and mini scale-up. The cost of the kit is less than the total cost of individual components.Catalysis Screening Kits... Read More | N665925 Component 1 mL 5 mL Storage N665925A 2×SYBR qPCR Master Mix 1 1 mL 5 mL -20℃. Avoid freeze/ Thaw cycle. N665925B qPCR Primer Mix 1 100 µL 500 µL -20℃. Avoid freeze/ Thaw cycle. N665925C DNA Standard I 100 µL 500 µL -20℃. Avoid freeze/ Thaw cycle. N665925 Component 1 mL 5 mL Storage N665925A 2×SYBR qPCR Master Mix 1 1 mL 5 mL -20℃. Avoid freeze/ Thaw cycle. N665925B qPCR Primer Mix 1 100 µL 500 µL -20℃. Avoid freeze/ Thaw cycle. N665925C DNA Standard I 100 µL 500 µL -20℃. Avoid freeze/ Thaw cycle. N665925D DNA Standard II 100 µL 500 µL -20℃. Avoid freeze/ Thaw cycle. N665925E DNA Standard III 100 µL 500 µL -20℃. Avoid freeze/ Thaw cycle. N665925F DNA Standard IV 100 µL 500 µL -20℃. Avoid freeze/ Thaw cycle. N665925G DNA Standard V 100 µL 500 µL -20℃. Avoid freeze/ Thaw cycle. N665925H 50×High ROX 40 µL 200 µL -20℃. Avoid freeze/ Thaw cycle.Product IntroductionThis product is a real-time fluorescence quantitative PCR of the products after NGS library construction using a dye method (SYBR Green I).(qPCR). The kit provides the reaction mixes, DNA primer mixtures, standards and sample dilutions required for the qPCR process, making the reagent system complete and easy to use. The fluorescent dye SYBR Green I contained in the reaction mixture can bind to all double-stranded DNA; the GoldStar Taq DNA Polymerase used is a chemically modified new high-efficiency hot-start polymerase, and the activation of the enzyme needs to be incubated at 95℃ for 10 minutes. the product has high specificity, high amplification efficiency, and is able to quickly and accurately quantify the concentration of the constructed libraries. The product is highly specific and efficient in amplification, and can quickly and accurately quantify the concentration of the constructed library.ROX dye is used to correct the fluorescence signal error generated between wells of a quantitative PCR instrument, and is generally used in Real Time PCR amplifiers from ABI, Stratagene, and other companies. The excitation optics vary from instrument to instrument, so the concentration of ROX dye must be matched to the corresponding fluorescence quantitative PCR instrument.Instruments that do not require ROX calibration: Roche LightCycler 480, Roche LightCyler 96, Bio-rad iCyler iQ, iQ5, CFX96, etc.Instruments requiring Low ROX calibration: ABI Prism7500/7500 Fast, QuantStudio®3 System, QuantStudio®5 System, QuantStudio®6 Flex System, QuantStudio®7 Flex System, ViiA 7 System, Stratagene Mx3000/Mx3005P, Corbett Rotor Gene 3000, and others.Instruments requiring High ROX calibration: ABI Prism7000/7300/7700/7900, Eppendorf, ABI Step One/Step One Plus, etc.Note: High Rox and Low Rox are formulated as described in Use 2.Scope of applicationThis product is designed for absolute quantification of the concentration of Ion torrent platform second generation sequencing libraries. The end of the library contains Ion torrent P5 and P7 microarray binding sequences, the length of which does not exceed 1kb, and the concentration is not less than 0.005pM can be used to perform quantitative experiments with this product. The qPCR Primer Mix provided in the kit contains the following two primer sequences:Primer 1:5'-CCA TCT CAT CCC TGC GTG TC - 3' Primer 2: 5'-CCT CTC TAT GGG CAG TCG GTG AT-3'The primer sequence can be used in advance to confirm whether the library can be amplified by that primer pair.Usage1. Amplification template preparationThe library samples to be detected were diluted with TE (10 mM Tris-Cl, pH 8.0, 1 mM EDTA), and the concentration after dilution was as close as possible to the range of 0.05-50 pM. 4°C on ice was set aside.2. qPCR reaction system preparationThe desired cryopreservation reagent is pre-melted completely and mixed by inverting several times before preparation, then centrifuged briefly and set aside.The base reaction system for 20 µl was as follows:Reagent20 µl Reaction system2×SYBR qPCR MasterMix10 µlqPCR Primer Mix 10.8 µlTemplate4 µlddH₂O5.2 µlDescription: High Rox model: add 1 µl High Rox per 50 µl of reaction system;Low Rox model: 1 µl High Rox per 500 µl of reaction system.Prepare a sufficient amount of reaction system mixture according to the need, mix well and add to the reaction wells in a volume of 16 µl per well, add the same volume of TE to the blank control, and then add the prepared standards and diluted samples to the corresponding reaction wells in a volume of 4 µl/well. It is recommended to use 20 µl reaction system, if you need to carry out a smaller system reaction, the system components can be reduced in equal proportion.3. qPCR reaction program1) Please use 60-64℃ as a reference for setting range of annealing temperature, and increase the annealing temperature when non-specific reaction occurs.2) If the average length of the library is greater than 700bp, the annealing/extension time should be increased appropriately.data analysis1. Standard curve productionThe standard curve was plotted using Ct values in the valid range. The standard curve correlation coefficient R2 should not be less than 0.99 and the slope should lie between -3.1 and -3.6. If the standard curve parameters are not reasonable, it is recommended to repeat the experiment.DNA Standard NameDNA Standard ConcentrationDNA Standard I50 pMDNA Standard II5 pMDNA Standard III0.5 pMDNA Standard IV0.05 pMDNA Standard V0.005 pM2. Library concentration calculationThe difference in Ct between the three replicate wells of the experiment should be no more than 0.2, otherwise the invalid data should be deleted or the experiment should be repeated. Do not use the Ct outside the valid Ct range of the standard curve to calculate the concentration of the diluted libraries. Please refer to the data processing Excel of this product for the specific library concentration calculation method.matters needing attention1. Before testing, these instructions should be read in detail. It should be operated by personnel with professional experience or qualified by training.2. For use, please mix gently by turning up and down, avoid foaming as much as possible, and use it after centrifugation for a short period of time.3. Avoid repeated freezing and thawing of the product, repeated freezing and thawing may degrade the performance of the product.4. When preparing the reaction solution, please use new or non-contaminated tips and centrifuge tubes to prevent contamination as much as possible... Read More | Product introduction：This kit uses an improved SDS alkaline lysis method combined with DNA preparation membrane to selectively adsorb DNA to achieve the purpose of rapid purification of plasmid DNA. It is suitable for extracting up to 500ug of high-purity plasmid DNA from 120-300ml bacterial Product introduction：This kit uses an improved SDS alkaline lysis method combined with DNA preparation membrane to selectively adsorb DNA to achieve the purpose of rapid purification of plasmid DNA. It is suitable for extracting up to 500ug of high-purity plasmid DNA from 120-300ml bacterial culture for sequencing, in vitro transcription and translation, restriction enzyme digestion, bacterial transformation and other molecular biology experiments.Scope of application：Nucleic acid extraction and purification... Read More | Products contentProducts IntroductionThis product uses the principle that the difference between the concentration of salt ions inside and outside the cell can cause the cell membrane to burst to lyses the cell and releases the genomic DNA, without the need of extracting and purifying the genomic Products contentProducts IntroductionThis product uses the principle that the difference between the concentration of salt ions inside and outside the cell can cause the cell membrane to burst to lyses the cell and releases the genomic DNA, without the need of extracting and purifying the genomic DNA.This product is suitable for a variety of sources of samples, and can be used as a template for PCR and qPCR experiments after sample processing, and can achieve the effect of the purified DNA used as a template for PCR and qPCR experiments. Usage1. Depending on the type of sample, prepare the appropriate sample size according to the table below.2. Add the sample to a 1.5-mi centrifuge tube and add the recommended volume of Solution A as shown in the table below. Vortex for 20 s and allow to stand at room temperature for 3-5 min or incubate in a metal bath at 95°C for 3-5 min as recommended in the table below.3. After the sample has been sufficiently lysed (samples incubated in a metal bath at 95°C should be brought to room temperature), add the recommended volume of Solution B as shown in the table below and vortex for 30s.4. Store processed samples at 4°C if the next test is to be performed within 2 hours, or at -20°C if the next test cannot be performed immediately.take note of1) Depending on the requirements of the experimental conditions, the amount of samples can be expanded or reduced, and the amount of Solution A and Solution B can be increased in equal proportions.2) For blood and cell samples, the temperature of room temperature lysis is required to be around 25C. If the ambient temperature does not reach 25°, the lysis time can be extended appropriately, or the vortex shaking time can be extended to ensure that the samples are fully lysed. If there is no relevant professional instrument, the centrifuge tube can be shaken vigorously to ensure adequate lysis.3) After the tissue sample is made into tissue homogenate by adding 10 times the volume of saline, it can be processed in the same way as blood samples.4) Strictly prohibit the use of expired products, please do not mix different reagents.5) laboratory supplies should be regularly cleaned and 10% of the 84 disinfectant solution or ultraviolet lamp for anti-pollution treatment, special areas dedicated to prohibit cross use, so as to avoid contamination, the end of the test, the bench should be cleaned immediately... Read More | Product contentY666144Component50 TStorageY666144ABuffer P115 mLRTY666144BBuffer P215 mLRTY666144CBuffer N320 mLRTY666144DBuffer PS15 mLRTY666144EBuffer PB10 mLRTY666144FBuffer PW (concentrate)10 mLRTY666144GBuffer EB10 mLRTY666144HGlass Beads2 gRTY666144IRNase A (10mg/mL)150 µLRTY666144JSpin Product contentY666144Component50 TStorageY666144ABuffer P115 mLRTY666144BBuffer P215 mLRTY666144CBuffer N320 mLRTY666144DBuffer PS15 mLRTY666144EBuffer PB10 mLRTY666144FBuffer PW (concentrate)10 mLRTY666144GBuffer EB10 mLRTY666144HGlass Beads2 gRTY666144IRNase A (10mg/mL)150 µLRTY666144JSpin Columns DM with Collection Tubes50 setsRTProductsThis kit is improved on the basis of common alkaline lysis method, the glass beads can effectively break the yeast cell wall, the new silica matrix membrane and buffer system can efficiently and specifically bind the plasmid DNA, and at the same time can maximize the removal of proteins and other impurities, the whole process is convenient and fast, no need to use toxic and harmful reagents, and can be processed at the same time for multiple samples. In addition to yeast cells, it can also be used in E. coli. Plasmid DNA extracted with this kit can be used in various molecular biology experiments, such as ligation, transformation, sequencing and library screening.Self-contained reagents: β-mercaptoethanol, anhydrous ethanol.Pre-experiment Preparation and Important Notes1. All components can be stably stored in dry, room temperature (15-30℃) environment for 1 year, the adsorption column can be stored at 2-8℃ for a longer period of time, and Buffer P1 with RNase A can be stably stored at 2-8℃ for 6 months.2. Before the first use, add all the RNase A solution to Buffer P1, mix well, and store at 2-8℃.3. Anhydrous ethanol should be added to Buffer PW before first use according to the instructions on the reagent bottle label.4. Before use, please check whether Buffer P2 and Buffer N3 are crystallized or precipitated. If there is any crystallization or precipitation phenomenon, it can be clarified by taking a water bath at 37℃ for a few minutes to restore the clarity.5. Be careful not to touch Buffer P2 and Buffer N3 directly, and tighten the lid immediately after use.6. The amount of plasmid extracted is related to the yeast strain, plasmid copy number, culture conditions, etc. Usually, yeast plasmid copy number is very low, which is difficult to be detected by electrophoresis or spectrophotometer method.Procedure1. Take 1-5 ml of yeast culture (maximum 5×107 yeast cells, generally for Saccharomyces cerevisiae OD = 1.0, equivalent to 1-2×107 cells/ml) and add it to a centrifuge tube (self-provided), centrifuge for 30 seconds at 12,000 rpm (~13,400×g), collect the bacterial precipitate, and aspirate as much as possible to discard the supernatant.2. Add 250µl Buffer P1 to the bacterium (please check if RNase A has been added first) and resuspend the precipitate.3. Add 40mg of Glass Beads to the above mixture and vortex and shake for 10 minutes.4. Add 250 µl of Buffer P2 to the centrifuge tube, mix gently by turning up and down 6-8 times, and let stand at room temperature for 5-10 minutes, at which time the bacterial solution should become clear and viscous.Note: Mix gently, do not shake violently, so as not to interrupt the genomic DNA, resulting in genomic DNA fragments mixed in the extracted plasmid. If the solution does not become clear, it suggests that the amount of bacteria may be too large and the lysis is not complete, and the amount of bacteria should be reduced.5. Add 350 µl of Buffer N3 to the centrifuge tube and immediately mix gently up and down 6-8 times, at which point a white flocculent precipitate appears, and centrifuge at 12,000 rpm for 20 minutes.Note: Buffer N3 should be mixed immediately after addition to avoid localized precipitation.6. Column Equilibration: Add 200 µl of Buffer PS to the Spin Columns DM in the collection tube, centrifuge at 12,000 rpm for 1 minute, pour off the waste liquid from the collection tube, and place the column back into the collection tube.7. Add the supernatant from step 5 to the adsorbent column that has been loaded into the collection tube, taking care not to aspirate the precipitate.Note: The maximum volume of the adsorption column is 750 µl, and the solution is passed through the column in 2 times.8. Centrifuge at 12,000 rpm for 1 minute, pour off the waste liquid in the collection tube and place the adsorption column back into the collection tube.9. Add 150 µl Buffer PB to the adsorbent column, centrifuge at 12,000 rpm for 1 min, pour off the waste liquid in the collection tube, and put the adsorbent column back into the collection tube.10. Add 750 µl Buffer PW to the adsorption column (please check that anhydrous ethanol has been added first), centrifuge at 12,000 rpm for 1 minute, and pour off the waste liquid in the collection tube.11. Place the column back into the recovery collection tube and centrifuge at 12,000 rpm for 2 minutes, pouring off the waste liquid. Leave the 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; ethanol residue can interfere with subsequent enzymatic reactions (digestion, PCR, etc.).12. Place the adsorbent column in a new centrifuge tube, add 50-100 µl of Buffer EB to the center of the adsorbent membrane dropwise, let it stand at room temperature for a few minutes, centrifuge at 13,000 rpm for 1 minute, and collect the plasmid solution into the centrifuge tube. Store the plasmid at -20°C.Attention:1) To increase the recovery efficiency of the plasmid, the resulting solution can be reintroduced into the adsorbent column, left at room temperature for a few minutes, centrifuged at 13,000 rpm for 1 minute, and the plasmid solution collected into a centrifuge tube.2) When the plasmid copy number is low or >10 kb, Buffer EB is preheated at 65-70°C in a water bath, which can increase the extraction efficiency.3) Usually yeast plasmids have very low copy number and are difficult to detect by electrophoresis or spectrophotometry. If the extracted plasmid is to be used in the next step of the experiment, it is usually recommended to use 1-5µl of the plasmid as PCR template, and 5-10µl of the plasmid for transformation of E. coli.4) Commercial high transformation efficiency receptor cells should be used for transformation of E. coli... Read More |