| Description | CTHRC1 Human Pre-designed siRNA Set A contains three designed siRNAs for CTHRC1 gene (Human), as well as a negative control, a positive control, and a FAM-labeled negative control. Components CTHRC1 siRNA-1: 5 nmol (HPLC) CTHRC1 siRNA-2: 5 nmol (HPLC) CTHRC1 siRNA-3: 5 nmol (HPLC) siRNA Negative CTHRC1 Human Pre-designed siRNA Set A contains three designed siRNAs for CTHRC1 gene (Human), as well as a negative control, a positive control, and a FAM-labeled negative control. Components CTHRC1 siRNA-1: 5 nmol (HPLC) CTHRC1 siRNA-2: 5 nmol (HPLC) CTHRC1 siRNA-3: 5 nmol (HPLC) siRNA Negative Control: 5 nmol (HPLC) FAM-labeled siRNA Negative Control: 5 nmol (HPLC) GAPDH siRNA Positive Control:5 nmol (HPLC)... Read More | Product introduction：Aladdin ® SE is a kind of fluorescent dye with amino reactive activity. The SE group of these dyes can react with the amino group to produce a stable amide bond. Compared with other similar dyes on the market, aladdin ® is a new generation of fluorescent dyes Product introduction：Aladdin ® SE is a kind of fluorescent dye with amino reactive activity. The SE group of these dyes can react with the amino group to produce a stable amide bond. Compared with other similar dyes on the market, aladdin ® is a new generation of fluorescent dyes with stronger stability, better water solubility and better fluorescence intensity. Product parameters： Absmax/Em(nm)：648/664；Absmax/Em(nm)：0.03；Extinction coefficient(ε)：240000；Optimal DOL(IgG)：3-6； Usage:1. Experimental materials(1) IgG: IgG must not contain amine chemicals that can react with dyes, such as amino acids, Tris, BSA, gelatin, etc. If IgG contains such chemicals, PBS buffer with pH~7.4 should be used for pre dialysis treatment. The presence of azide compounds does not affect the labeling reaction.(2) Anhydrous DMSO(3) NaHCO3(4) Sephadex gel G-25 dialysis column(5) PBS buffer (pH~7.4)(6) NaN3(7) BSA2. Marking methods and steps(1) Prepare to label antibodiesDilute the antibody with 0.1 M NaHCO3 solution (pH~8.3) to a final concentration of 2.5 mg/mL. If the product is pre diluted with phosphate buffer, such as PBS buffer (without amino compounds), approximately 1/10 volume of 1M NaHCO3 mother liquor can be directly added to the buffer to achieve a final NaHCO3 concentration of 0.1 M.Note: When the protein concentration is 2.5 mg/mL, the labeling efficiency is approximately 35%. Protein concentrations below 2.5 mg/mL can also be used for labeling, but the labeling efficiency will decrease. When the protein concentration is higher than 5 mg/mL, the labeling efficiency may be higher. Due to differences in buffer and protein purity, more precise labeling efficiency is determined by practical operating conditions. If the protein concentration is too low, it can be concentrated by ultrafiltration.(2) Prepare dye storage solutionPreheat one tube at room temperature µ YF of Mole ® SE, add 0.1 mL of anhydrous DMSO to the tube, thoroughly vortex dissolve the dye, and prepare a dye storage solution with a concentration of 10 mM. If a trace amount of protein is used for labeling reactions, the dye needs to be diluted to a lower concentration.Note: a The remaining dye storage solution should be stored at a low temperature of -20 ℃ for future use. If anhydrous DMSO is used to prepare dye storage solution, the dye can be stored for at least one month.b. Dyes can also be prepared with deionized water, but due to the slow hydrolysis of dyes in water, it is best to prepare water based storage solutions for immediate use.(3) Mark reaction stepsa. Stir or vortex the protein solution, gradually adding 15-25 drops µ L dye storage solution (10 mM), with a molar ratio of dye/protein in the range of 9:1 to 15:1. YF ® Please refer to the table above for the range of DOL (number of dyes bound to each protein molecule) for SE labeled IgG antibodies.b. Stir the reaction at room temperature for 1 hour, and for trace labeling, shake and incubate on a shaker for 1 hour.Note: At the same time of the binding reaction, proceed to step 2 (4) to balance the dextran gel G-25 dialysis column.(4) Isolation of marker proteins from reaction solutiona. PBS buffer (pH~7.4) was used to balance the dextran gel G-25 dialysis column (10 mm × 300 mm).b. Add the reaction solution from step 3 (b) to the column and elute with 1 x PBS buffer.The first washed out chromophore is a dye protein complex.Note: a For small-scale labeling reactions, in order to avoid excessive dilution of the product, ultrafiltration devices can be used to remove free dyes from the complex.b. After the binding reaction is completed, if the dye protein complex is not separated in time, 50 can be added µ Terminate the reaction with L 1M lysine. In most cases, this operation is not necessary because the remaining unreacted dyes have been fully hydrolyzed at the end of the reaction.3. Determine DOL(1) The determination of protein concentration and antibody concentration can be calculated using the following formula:C (mg/mL)={[A280- (Amax x x Cf)]/1.4} x dilution factor;a. C refers to the concentration of antibodies collected in the experiment;b. Dilution factor refers to the dilution factor used in photometric measurements;c. A280 and Amax refer to the absorbance at 280 nm and the absorbance at the absorption wavelength, respectively;d. Cf is the correction factor, YF ® Please refer to the table above for the Cf value of SE dyes;Note: The protein solution eluted through the column may have a high concentration when used directly for absorbance detection, so it needs to be diluted to approximately 0.1 mg/mL. The dilution factor (i.e. dilution factor) needs to be determined from the initial number of antibodies (e.g. 5 mg) and the overall elution of protein solutionEstimate based on the product.(2) Estimation of DOLDOL is calculated using the following equation:DOL=(Amax x x Mwt x Dilution Factor)/（ ε X C)a. Amax, dilution factor, C value has been clearly defined in 3 (1);b. Mwt refers to the molecular weight of IgG (150000);C. c ε It's YF ® The molar absorption coefficient of SE, refer to the table on the first page;d. Mark YF ® The optimal DOL value for SE IgG antibodies can be found in the table on the first page. Although DOL values may fluctuate, good experimental results can also be achieved.Matters needing attention：1. if the labeled protein needs long-term storage, it is recommended to add 5-10 mg/ml BSA and 0.01-0.03% NaN3 to prevent protein denaturation and microbial breeding. Store at 4 ℃ away from light. If glycerol with a final concentration of 50% is added, it can be stored at -20 ℃. It can be stably stored for more than one year. 2. keep away from light during operation. The mixing speed should be appropriate to avoid bubbles. 3. when installing the chromatographic column, try to make the column body uniform, the column surface flat, and free of bubbles and cracks. 4. pay attention to adding the sample when the column top buffer is tangent to the gel plane. When eluting, add the eluent when the sample is tangent to the gel plane. 5. other factors affecting the labeling efficiency also include temperature, reaction time, pH, the amount of fluorescent dye and protein, etc., which should be controlled. 6. for your safety and health, please wear laboratory clothes and disposable gloves.Scope of application：Protein nucleic acid labeling dye... Read More | This product is a mixture of fast reverse transcription reagents. The 5 x EasyQuick RT MasterMix contains all the reagents required for reverse transcription from RNA templates to cDNA first strand, including EasyQuick RT Reversase, RNase Inhibitor, Random 6 mers, Oligo dT Primer, dNTP, EQ-RT BufferThis product is a mixture of fast reverse transcription reagents. The 5 x EasyQuick RT MasterMix contains all the reagents required for reverse transcription from RNA templates to cDNA first strand, including EasyQuick RT Reversase, RNase Inhibitor, Random 6 mers, Oligo dT Primer, dNTP, EQ-RT Buffer, etc. The reverse transcription efficiency of this product is high, and it can perform a good reverse transcription reaction on a small amount of RNA templates. The fluorescence quantitative template cDNA first strand synthesis can be completed in 15 minutes. This reagent kit is very convenient and fast to operate, and only RNA templates and water need to be added for reverse transcription reaction, making it particularly suitable for high-throughput detection.E665905Component200 TStorageE665905A5×EasyQuick RT MasterMix 400 µL-20℃. Avoid freeze/thaw cycle.E665905BRNase-Free Water 2×1 mL-20℃. Avoid freeze/thaw cycle. Product features1. Convenience: The ready to use reverse transcription Mix only requires the addition of RNA templates and water to initiate the reaction.2. Fast: Complete cDNA first strand synthesis in 15 minutes.3. High reverse transcription efficiency: The reverse transcription efficiency is above 90%.4. High sensitivity: PG level templates can also obtain high-quality cDNA.5. Read through complex templates: templates with high GC content and complex secondary structures.Matters needing attention1. During the operation, RNase contamination should be avoided to prevent RNA degradation or cross contamination during experiments. It is recommended that operators wear masks and disposable gloves, frequently change gloves, and use specialized instruments and consumables.2. The reverse transcription system is prepared on ice for operation to prevent RNA degradation. The MasterMix of the reagent kit should be stored at -20 ℃ as soon as possible after use, and repeated freeze-thaw should be avoided as much as possible.3.10 µ The reaction system can be used up to 1 µ G Total RNA, if the amount of template RNA is greater than 1 µ g. Please expand the reaction system proportionally.4. For RNA templates with complex secondary structures, it is recommended to incubate the template RNA at 65 ℃ for 5 minutes on ice before proceeding with the next step, followed by brief centrifugation.Operation steps1. Thaw the template RNA on ice; After thawing the components of the reagent kit at room temperature, immediately place them on ice. Before use, vortex shake and mix each solution, and centrifuge briefly before use.2. Prepare the reaction system according to the following table (please prepare the reaction solution on ice), vortex shake and mix well, briefly centrifuge, and collect the solution on the tube wall to the bottom of the tube. Reagent 10 µl Reaction system Final concentration RNA Template X µl 1 pg～0.5 µg ¹⁾ 5×EasyQuick RT MasterMix ²⁾ 2 µl 1× RNase-Free Water up to 10 µl /Attention:1) If the total RNA content is greater than 1 µ g, please expand the reaction system proportionally.2) 5 x EasyQuick RT MasterMix contains Oligo (dT), Random Prime, RNase Inhibitor, dNTP Mixture, EQ-RT Buffer, etc. 3. Incubate at 37 ℃ for 15 minutes. 4. Incubate at 85 ℃ for 5 seconds to inactivate reverse transcriptase.5. After a brief centrifugation, place it on ice for subsequent experiments. If it needs to be stored for a long time, please place it at -20 ℃... Read More | Product contentF665766Component5 mLStorageF665766A2×Fast Probe Mixture5×1 mL-20℃. Avoid freeze/thaw cycle.F665766BddH2O5×1 mL-20℃. Avoid freeze/thaw cycle.Product IntroductionFast Probe Mixture is a pre-mixed system for real-time fluorescence PCR by probe method (TaqMan, Product contentF665766Component5 mLStorageF665766A2×Fast Probe Mixture5×1 mL-20℃. Avoid freeze/thaw cycle.F665766BddH2O5×1 mL-20℃. Avoid freeze/thaw cycle.Product IntroductionFast Probe Mixture is a pre-mixed system for real-time fluorescence PCR by probe method (TaqMan, Molecular Beacon, etc.), with a concentration of 2×, including Fast Taq DNA Polymerase, PCR Buffer, dNTPs, Mg2+ and so on, which is easy and convenient to operate. It is mainly used for the detection of genomic DNA target sequence and cDNA target sequence after RNA reverse transcription. The Fast Taq DNA Polymerase contained in this product can effectively reduce the non-specific amplification generated by the non-specific binding of primers and templates or primer dimerization at room temperature, and the activation of the enzyme only needs to be incubated at 95 ℃ for 30 s. The whole PCR reaction process can save about 40 minutes compared with the ordinary reaction, which greatly shortens the reaction time of PCR. The combination of unique PCR buffer system and fast hot start enzyme effectively inhibits the generation of non-specific products and significantly improves the PCR amplification efficiency with stronger fluorescence signal, higher sensitivity and wider linear range. The product has a wide range of applications and can be used for both normal and rapid quantitative PCR programs.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 (F665766):Roche LightCycler 480, Roche LightCyler 96, Bio-rad iCyler iQ, iQ5, CFX96 and others.Instruments that require Low ROX calibration (F665768):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 more.Instruments that require High ROX calibration (F665774):ABI Prism 7000/7300/7700/7900, Eppendorf, ABI Step One/Step One Plus, and others.matters needing attention1. Before use, please mix gently by turning up and down, avoid foaming as much as possible, and use after brief centrifugation.2. Avoid repeated freezing and thawing of this product, repeated freezing and thawing may degrade the product performance. This product can be stored for long term at -20℃, protected from light. If frequent use is required within a short period of time, it can be stored at 2-8℃.UsageThe following examples are conventional PCR reaction systems and reaction conditions, which should be improved and optimized according to the template, primer structure and target fragment size in actual operation.1.PCR reaction systemReagents50 µl Reaction systemFinal concentration2×Fast Probe Mixture25 µl1×Forward Primer，10 µM1 µl0.2 µM¹⁾Reverse Primer，10 µM1 µl0.2 µM¹⁾Probe，10 µM1 µl0.2 µM²⁾Template DNA2 µl³⁾ 50×Low ROX or High ROX（optional）⁴⁾1 µl1×ddH₂Oup to 50 µlNote: 1) Usually the primer concentration of 0.2µM can get better results, and 0.1-1.0µM can be used as a reference for setting the range. 2) The final concentration of the probe used is related to the fluorescent quantitative PCR instrument used, the type of probe, and the type of fluorescent labeling substance, so please refer to the instruction manual of the instrument or the specific requirements of the use of each fluorescent probe for the adjustment of the concentration in actual use.(3) Usually the amount of DNA template is 10-100ng genomic DNA or 1-10ng cDNA as a reference. Since the templates of different species contain different copy numbers of target genes, the templates can be subjected to gradient dilution to determine the optimal amount of template to be used.(4) The excitation optical system varies from instrument to instrument, choose to add 50×Low ROX or 50×High ROX according to the instrument using fluorescence quantification.2. PCR reaction program:A two-step PCR reaction program is recommended, and this program is set up using the ABI 7500 Fluorescent Quantitative PCR Instrument as a reference.Note: 1) The enzyme used in this product must be pre-denatured at 95°C for 30s to achieve enzyme activation. Under this condition, most of the templates can be well unchained. For templates with high GC content and complex secondary structure, the pre-denaturation time can be extended to 1-4 minutes in order to make the starting template fully unchained.(2) It is recommended to use two-step PCR reaction program, if you can not get good experimental results due to the use of primers with lower Tm values, etc., you can try to carry out a three-step PCR amplification, annealing temperature, please use the range of 56 ℃ - 64 ℃ for as a reference for the setting... Read More | Reverse transcriptases are enzymes encoded in retroviruses viral genome. The enzyme is responsible for transcription of the viral RNA to produce a dsDNA that can be inserted into the host genome.Reverse transcriptases are multifunctional enzymes. These enzymes exhibit an RNA and DNA directed Reverse transcriptases are enzymes encoded in retroviruses viral genome. The enzyme is responsible for transcription of the viral RNA to produce a dsDNA that can be inserted into the host genome.Reverse transcriptases are multifunctional enzymes. These enzymes exhibit an RNA and DNA directed polymerase activity. In addition reverse transcriptases catalyze the degradation of RNA in an RNA-DNA hybrid. The exonucleolytic activity proceeds in a 5' ---> 3' direction. The RNA or DNA directed activity requires a template (RNA or DNA) and a primer. The following is a schematic illustration of the reaction:Unit definition: One unit incorporates 1 nanomole of tritiated dTMP into acid insoluble productsusing poly(A)•oligo(dT) 12-18 as the template-primer in 20 minutes at 37° C.ApplicationsHIV reverse transcriptase is used for research on the AIDS primer. However it can be substituted for AMV reverse transcriptase, which is mainly used to transcribe mRNA into double stranded cDNA, that can be inserted into prokaryotic vectors. The enzyme can also be used with either single stranded DNA or RNA templates to make probes for use in hybridization experiments. It can be used for labeling the termini of DNA fragments with protruding 5' termini. The enzyme can also be used to sequence DNAs by the dideoxy chain termination method of Sanger when the Klenow fragment of E. coli DNA polymerase I, or the T7 DNA polymerase yield unsatisfactory results.Reagents0.05 M Tris, pH 8.3, containing 0.008 M MgCl21 mg/ml polyadenylic acid in water (poly A)DNA primer:Oligo d(T)12-181 µ mole dTTP/mL stock solution[methyl-3H]-Thymidine 5'-triphosphate (3H-dTTP)dTTP-3H-dTTP working mix: Add 1-2 µL 3H-dTTP per mL of 100 nmol/mL dTTP in order to obtain 1 to 1.5 x 105 cpm/mL1% bovine serum albumin10% perchloric acid1% perchloric acidBuffer substrate reaction mixture: Prepare fresh, immediately before use:For each 1mL of reaction mixture required mix:0.7 mL Tris/HCl, pH 8.3, 0.008M MgCl20.3 mL 1 mg/mL poly(A) RNA template0.005 mL 0.02 mg/mL oligo d(T)12-18 DNA primer0.02mL 1% BSAEnzymedilute as needed wtih 0.05M Tris/HCl, pH 8.3, 0.008M MgCl2 containing 0.1 mg/mL (1%) BSAProcedurePipette into each tube as follows:Buffer substrate mix：0.1 mLdTTP-3H3-dTTP：0.1 mLEnzyme：5-10 µLIncubate 20 minutes at 37° C. Stop reaction by adding 1 ml 10% cold perchloric acid. Filter through 0.2µ manifold filters used with Millipore vacuum manifold. Wash four times using 2mL 1% cold perchloric acid/wash. Transfer filter to scintillation vials. Add 2mL Cellosolve (or 2-methoxyethanol) to dissolve filter. Filters become opaque upon addition of Cellosolve. Make sure filters are dissolved before proceeding. Add 10mL scintillation cocktail and count.Calculation... Read More |