| Description | Hotstart HiTaq& Super M-MuLV One Step RT-qPCR Kit (with UDG) is widely recognized for its exceptional speed and quantitative accuracy. This product enables continuous Real-Time RT-PCR reactions within a single tube, simplifying operations. The enzyme system employs thermolabile UDG (Uracil-DNA Hotstart HiTaq& Super M-MuLV One Step RT-qPCR Kit (with UDG) is widely recognized for its exceptional speed and quantitative accuracy. This product enables continuous Real-Time RT-PCR reactions within a single tube, simplifying operations. The enzyme system employs thermolabile UDG (Uracil-DNA Glycosylase) in combination with dUTP-containing substrates, effectively preventing carryover contamination from previous PCR products. The thermolabile UDG is completely inactivated at 50°C, ensuring it does not interfere with the reverse transcription products of the current reaction. This kit utilizes a highly stable and efficient reverse transcriptase (Super M-MuLV Reverse Transcriptase) and a chemically modified hot-start DNA polymerase (Hotstart HiTaq DNA Polymerase). It delivers high amplification efficiency and specificity, enabling robust Real-Time One-Step RT-PCR. This significantly enhances detection sensitivity and eliminates the need for post-PCR electrophoresis, making it particularly suitable for the detection of trace amounts of RNA. Specifically designed for convenient and highly sensitive one-step RT-PCR, this kit features a unique enzyme formulation and specially optimized buffer system. These ensure efficient and accurate reverse transcription and PCR amplification without the need for reaction optimization. It generates excellent standard curves over a wide quantitative range, allowing precise quantitative detection of target genes with high reproducibility and reliability. Applications: Quantitative or qualitative detection of nucleic acids using RNA as template. Reverse transcription and amplification are completed in a single tube without tube opening.Product Component ListU1492590Component100T1000T10000TStorageU1492590AHotstart HiTaq& Super M-MuLV Mix0.2 mL2 mL20 mL-20℃U1492590BBuffer Mix0.95 mL9.5 mL95 mL-20℃ Protocol:1. Reaction Setup:Thaw and thoroughly mix all PCR solutions required for the reaction.Briefly centrifuge the solutions and keep them at room temperature until use.Probes should be protected from light.Recommendation: Aliquot PCR reagents to avoid repeated freeze-thaw cycles.2. Prepare the PCR Reaction Mixture according to the table below:ComponentVolume/25 µLVolume/50 µLFinal ConcentrationBuffer Mix9.5 µL19 µL1×Enzyme Mix (Hotstart HiTaq& Super M-MuLV)2 µL4 µL2/50 µLPrimer-Probe Mix1 µL2 µL/Template5 µL10 µL/ddH2OTo 25 µLTo 50 µL/Total Volume25 µL50 µLNote: The amounts of primers, probes, and template can be adjusted according to experimental requirements.3. Mixing:Gently mix the reaction components by pipetting up and down or by brief vortexing. Centrifuge briefly at room temperature to collect the liquid at the bottom of the tube.4. PCR Amplification:Place the PCR reaction tubes in the thermal cycler and initiate the PCR program.Standard Thermal Cycling Protocol:StepTemperatureTimeCyclesReverse Transcription50℃15min1Initial Denaturation95℃10min1Denaturation95℃10s45Annealing/Extension60℃40s45Hold12℃∞1Note: Reaction conditions can be adjusted based on specific experimental requirements... Read More | Product content:D665967Component200 TStorageD665967ABuffer PB120 mLRTD665967BBuffer PS60 mLRTD665967CBuffer PW (concentrate)25 mLRTD665967DBuffer EB30 mLRTD665967ESpin Columns DM with Collection Tubes200 EART Product Introduction: This reagent kit adopts a new silicon-based membrane technology and Product content:D665967Component200 TStorageD665967ABuffer PB120 mLRTD665967BBuffer PS60 mLRTD665967CBuffer PW (concentrate)25 mLRTD665967DBuffer EB30 mLRTD665967ESpin Columns DM with Collection Tubes200 EART Product Introduction: This reagent kit adopts a new silicon-based membrane technology and reagent formula. Through a rapid and simple three-step process of binding, washing, and elution, 100 bp-10 kb DNA fragments can be purified and recovered from PCR products or enzyme reaction solutions (enzyme cutting, linking, probe labeling, etc.). Each adsorption column can adsorb up to 10 kb of DNA fragments µ G DNA, while minimizing impurities such as primers, oligonucleotides, enzymes, etc. The purified and recovered DNA has high purity and concentration, good integrity, and high recovery rate, and can be directly used for molecular biology experiments such as sequencing, linking and transformation, labeling, and in vitro transcription.Self prepared reagent: anhydrous ethanol.Preparation and important precautions before the experiment:1. All components can be stably stored in a dry, room temperature (15-30 ℃) environment for 1 year, and can be stored at 2-8 ℃ for longer periods of time. When the solution is stored at low temperature, it should be left at room temperature for a period of time before use, and then restored to room temperature before use.2. This reagent kit can selectively recover all DNA fragments from the solution. If you need to selectively recover specific fragments while removing other fragments of different sizes, please choose our company's gel recovery reagent kit.3.Before the first use, anhydrous ethanol should be added to the Buffer PW according to the instructions on the reagent bottle label.4. Before use, please check if there is any crystallization or precipitation in the Buffer PB. If there is any crystallization or precipitation, you can take a water bath at 37 ℃ for a few minutes to restore clarity.5. The recovery efficiency is related to the initial amount of DNA and the elution volume. The smaller the initial amount, the smaller the elution volume, and the lower the recovery rate.6. All centrifugation steps can be performed at room temperature.Operation steps:1. Estimate the volume of DNA reaction solution, add 5 times the volume of Buffer PB, and mix thoroughly (without removing paraffin or mineral oil).Note: 1) If the DNA reaction system is 50 µ l (excluding paraffin oil volume), add 250 µ l Buffer PB.2) After adding Buffer PB, check the pH value of the solution. If the pH value is greater than 7.5, add 10-30 to it µ 3 M sodium acetate (pH 5.0) was used to adjust the pH value to 5-7.2. Column balance: Add 200 to the spin columns DM that have been loaded into the collection tube µ Centrifuge at 13000 rpm (~16200 × g) for 1 minute, discard the waste liquid in the collection tube, and place the adsorption column back into the collection tube.3. Add the solution obtained in step 1 to the adsorption column that has been loaded into the collection tube, let it stand at room temperature for 1 minute, centrifuge at 13000 rpm for 30-60 seconds, discard the waste liquid in the collection tube, and place the adsorption column in the collection tube.Attention: The volume of the adsorption column is 750 µ l. If the sample volume is greater than 750 µ l, it can be added in batches.4. Add 500 µ l of Buffer PW to the adsorption column (please check if anhydrous ethanol has been added before use), centrifuge at 13000 rpm for 30-60 seconds, discard the waste liquid in the collection tube, and place the adsorption column in the recovery tube.Note: If purified DNA is used for salt sensitivity experiments (such as flat end ligation experiments or direct sequencing), it is recommended to add Buffer PW and let it stand for 2-5 minutes before centrifugation.5.13000 rpm for 1 minute and discard the waste liquid from the collection tube. Place the adsorption column at room temperature for a few minutes to thoroughly air dry.Note: The purpose of this step is to remove residual ethanol from the adsorption column, which can affect subsequent enzymatic reactions (such as enzyme digestion, PCR, etc.). To ensure that downstream experiments are not affected by residual ethanol, it is recommended to open the cover of the adsorption column and place it at room temperature for a few minutes to thoroughly dry the residual ethanol in the adsorbent material at the bottom.6. Place the adsorption column into a new centrifuge tube (provided by oneself), add 30-50 µ l Buffer EB to the middle position of the adsorption membrane by hanging droplets, and let it stand at room temperature for 1 minute. Centrifuge at 13000 rpm for 1 minute and collect DNA solution- Store DNA at 20 ℃.Attention:1) The pH value of the eluent has a significant impact on the elution efficiency. If water is used as the eluent, its pH value should be ensured to be between 7.0-8.5 (the pH value of water can be adjusted to this range using NaOH).2) To improve the recovery of DNA, the solution obtained by centrifugation can be added back to the adsorption column, left at room temperature for 2 minutes, and centrifuged at 13000 rpm for 1 minute.3) The elution volume should not be less than 30 µ l. A small volume will affect the recovery efficiency... Read More | M666110 Component 96 T Storage M666110A Buffer WSL 40 mL RT M666110B Buffer MSL 40 mL RT M666110C Buffer CW1 (concentrate) 90 mL RT M666110D Buffer GW1 (concentrate) 40 mL RT M666110E Buffer GW2 (concentrate) 50 mL RT M666110F Buffer EB 30 mL RT M666110G Proteinase K 4×1.25 mL RT M666110H M666110 Component 96 T Storage M666110A Buffer WSL 40 mL RT M666110B Buffer MSL 40 mL RT M666110C Buffer CW1 (concentrate) 90 mL RT M666110D Buffer GW1 (concentrate) 40 mL RT M666110E Buffer GW2 (concentrate) 50 mL RT M666110F Buffer EB 30 mL RT M666110G Proteinase K 4×1.25 mL RT M666110H Magbeads V3 2×1 mL RTProduct Introduction：The reagent kit provides a simple, fast, and efficient method for extracting genomic DNA from blood samples. In the presence of high salt, DNA binds to the surface of silica coated Magheads. After rinsing, high-purity DNA is eluted in Buffer EB or deionized water. The purified DNA has good purity (A260/280 ratio between 1.7-1.9) and high integrity (>15 kb), and can be used for downstream experiments such as second-generation sequencing, quantitative PCR, and chip detection.Self provided instruments and reagents1) Constant temperature mixer2) 2/15 ml magnetic frame3) 32 channel nucleic acid extractor4) 96 channel nucleic acid extractor5) 96 DW Plate6) 8 channel Comb7) Spin tips pack8) Anhydrous ethanolPreparation and important precautions before the experiment1.Before the first use, add anhydrous ethanol to Buffer CW1, Buffer GW1, and Buffer GW2 according to the label of the reagent bottle and mark them properly.2.Magheads are strictly prohibited from freezing or centrifugation. Freezing and centrifugation may cause irreversible damage to Magheads.Operation stepsI. Manual single tube operation1. Use punching forceps to take 1 blood spot with a diameter of 6 mm or 4 blood spots with a diameter of 3 mm (depending on the actual situation) from the blood spot and place them in a 2.0 mL centrifuge tube.2. Add 40 to the centrifuge tube µ L Protein K and 300 µ L Buffer WSL, then place the centrifuge tube on a constant temperature mixer at 75 ℃ and 1200 rpm, shake and crack for 45 minutes to form Lysate. Remove the centrifuge tube from the constant temperature mixer, centrifuge briefly, and take the supernatant.Attention: If there is no constant temperature mixer, vortex the centrifuge tube for 10 seconds and incubate it in a 75 ℃ water bath for 30 minutes. During this period, vortex every 10 minutes for 10 seconds.3. Suck the supernatant into a new 2.0 mL centrifuge tube and add 300 µ L Buffer MSL, 300 µ L isopropanol and 20 µ L Magheads V3. Afterwards, place the centrifuge tube on a constant temperature mixer at 25 ℃ and 1600 rpm, shake and crack for 15 minutes, or invert the centrifuge tube and mix continuously for 15 minutes.4. Place the centrifuge tube on a magnetic stand and let it stand for 1 minute. After Magheads are completely adsorbed on the side wall of the centrifuge tube, discard the solution thoroughly (keep the centrifuge tube fixed on the magnetic stand).5. Remove the centrifuge tube from the magnetic frame and add 900 µ L Buffer CW1 (please check if anhydrous ethanol has been added before use), vortex point shake for 1 minute or vortex shake for 5 seconds, and then place it on a constant temperature mixer at 25 ℃ and 1600 rpm to shake and mix for 2 minutes (ensure that Magheads are in a mixed state during the shaking process). Afterwards, place the centrifuge tube on a magnetic stand and let it stand for 1 minute. After Magheads are completely adsorbed on the side wall of the centrifuge tube, gently invert the magnetic stand and wash the impurities on the centrifuge tube cover to completely discard the solution (keep the centrifuge tube fixed on the magnetic stand).6. Remove the centrifuge tube from the magnetic frame and add 500 µ L Buffer GW1 (please check if anhydrous ethanol has been added before use), vortex point shake for 1 minute or vortex shake for 5 seconds, and then place it on a constant temperature mixer at 25 ℃ and 1600 rpm to shake and mix for 2 minutes (ensure that Magheads are in a mixed state during the shaking process). Afterwards, place the centrifuge tube on a magnetic stand and let it stand for 1 minute. After Magheads are completely adsorbed on the side wall of the centrifuge tube, gently invert the magnetic stand and wash the impurities on the centrifuge tube cover to completely discard the solution (keep the centrifuge tube fixed on the magnetic stand).7. Remove the centrifuge tube from the magnetic frame and add 900 µ L Buffer GW2 (please check if anhydrous ethanol has been added before use), vortex point shake for 1 minute or vortex shake for 5 seconds, then place it on a constant temperature mixer at 25 ℃ and 1600 rpm, shake and mix for 2 minutes (ensure that Magheads are in a mixed state during the shaking process). Afterwards, place the centrifuge tube on a magnetic stand and let it stand for 1 minute. After Magheads are completely adsorbed on the side wall of the centrifuge tube, gently invert the magnetic stand and wash the impurities on the centrifuge tube cover to completely discard the solution (keep the centrifuge tube fixed on the magnetic stand).8. Remove the centrifuge tube from the magnetic frame and add 300 µ After shaking with 75% ethanol for 1 minute or 5 seconds, place the mixture on a constant temperature mixer at 25 ℃ and 1600 rpm for 2 minutes (ensure that the Magheads are in a mixed state during the shaking process). Afterwards, place the centrifuge tube on a magnetic stand and let it stand for 1 minute. After Magheads are completely adsorbed on the side wall of the centrifuge tube, gently invert the magnetic stand and wash the impurities on the centrifuge tube cover to completely discard the solution (keep the centrifuge tube fixed on the magnetic stand).9. Keep the centrifuge tube fixed on the magnetic frame, use a pipette to further remove the solution from the bottom and cover of the centrifuge tube, and then leave it at room temperature for 5-10 minutes to allow the ethanol to evaporate completely.10. Remove the centrifuge tube from the magnetic frame and add 50-200 µ L Buffer EB. Vortex oscillation causes the magnetic beads to completely suspend in the eluent and then place them on a constant temperature mixer at 56 ℃ and 1600 rpm for 10 minutes of shaking and elution, or incubate the centrifuge tube in a 56 ℃ water bath for 10 minutes, with vortex oscillation every 3 minutes for 10 seconds.11. Place the centrifuge tube on a magnetic stand and let it stand for 2 minutes. After Magheads are completely adsorbed on the side wall of the centrifuge tube, transfer the eluent to a new centrifuge tube using a pipette and store at -20 ℃ for later use.II. Matching with CWE21001. Use punching forceps to take 1 blood spot with a diameter of 6 mm or 4 blood spots with a diameter of 3 mm (depending on the actual situation) from the blood spot and place them in a 2.0 mL centrifuge tube.2. Add 40 to the centrifuge tube µ L Protein K and 300 µ L Buffer WSL, then place the centrifuge tube on a constant temperature mixer at 75 ℃ and 1200 rpm, shake and crack for 45 minutes to form Lysate.3. Add the corresponding reagents to the 96DW deep well plate according to the table below. Position Reagent 1&7 Colume Lysate: All Buffer MSL: 300 µL isopropanol：300 µL Magbeads V3: 20 µL 2&8 Colume Buffer CW1: 900 µL 3&9 Colume Buffer GW1: 500 µL 4& 10 Colume Buffer GW2: 900 µL 5& 11 Colume 75%ethanol: 300 µL 6& 12 Colume Buffer EB: 70 µL4.Place the deep well plate and magnetic sleeve that have been added to the reagent at the corresponding positions of CWE2100/CWE3200, run the blood slide extraction program, and after about 40 minutes, the program ends. Remove the deep well plate and magnetic sleeve.5.Transfer the elution products from columns 6 and 12 of the deep well plate to a 1.5 mL centrifuge tube for low-temperature storage.III. Matching with CWE9601. Use punching forceps to take 1 blood spot with a diameter of 6 mm or 4 blood spots with a diameter of 3 mm (depending on the actual situation) from the blood spot and place them in a 2.0 mL centrifuge tube.2. Add 40 to the centrifuge tube µ L Protein K and 300 µ L Buffer WSL, then place the centrifuge tube on a constant temperature mixer at 75 ℃ and 1200 rpm, shake and crack for 45 minutes to form Lysate.3. Add the corresponding reagents to the 96DW deep well plate according to the table below Position Reagent Plate 1 Lysate: All Buffer MSL: 300 µL isopropanol ：300 µL Magbeads V3: 20 µL Plate 2 Buffer CW1: 900 µL Plate 3 Buffer GW1: 500 µL Plate 4 Buffer GW2: 900 µL Plate 5 75% ethanol : 300 µL Plate 6 Buffer EB: 70 µL4. Place the deep well plate and magnetic sleeve that have been added to the reagent at the corresponding positions on CWE960, run the blood slide extraction program, and after about 40 minutes, the program ends. Remove the deep well plate and magnetic sleeve.5. Transfer the elution products from Plate 6 to a 1.5 mL centrifuge tube for low-temperature storage... Read More | The content of this cell is too long for an XLSX file (more than 32767 characters). Please use the CSV format for this export | Product contentS665868Component50 TStorageS665868ABuffer GL25 mLRTS665868BBuffer GW1 (concentrate)13 mLRTS665868CBuffer GW2 (concentrate)15 mLRTS665868DBuffer GE15 mLRTS665868EProteinase K2×1.25 mLRTS665868FSpin Columns DM with Collection Tubes50 setsRTProduct IntroductionThis kit is suitable Product contentS665868Component50 TStorageS665868ABuffer GL25 mLRTS665868BBuffer GW1 (concentrate)13 mLRTS665868CBuffer GW2 (concentrate)15 mLRTS665868DBuffer GE15 mLRTS665868EProteinase K2×1.25 mLRTS665868FSpin Columns DM with Collection Tubes50 setsRTProduct IntroductionThis kit is suitable for the extraction of genomic DNA from fresh saliva or saliva/preservation solution mixture.The purification process of this product does not require the use of toxic solvents such as phenol or chloroform, and ethanol precipitation is not necessary. The optimized buffer system enables DNA to bind heterogeneously to the silica matrix centrifugal adsorption column, and the inhibitors of PCR and other enzymatic reactions can be effectively removed by a two-step washing step, and finally eluted with a low-salt buffer or water to obtain high-purity DNA.The purified obtained can be directly used for enzyme digestion, PCR, Real-Time PCR, library construction, Southern Blot, molecular labeling and other downstream experiments.Self-contained reagent: anhydrous ethanolPre-experiment Preparation and Important Notes1. Repeated freezing and thawing of the sample should be avoided, as this may result in smaller fragments of extracted DNA and a decrease in the amount extracted.2. Anhydrous ethanol should be added to Buffer GW1 and Buffer GW2 according to the instructions on the label of the reagent bottle before first use.3. Before use, please check whether Buffer GL appears to be crystallized or precipitated.Redissolve in a 56°C water bath.4. If the downstream experiments are sensitive to RNA contamination, 4 µL DNase-Free RNase A can be added in step 3(100 mg/mL).5. For prolonged storage of salivary DNA at room temperature, our Salivary DNA Preservation Tubes are recommended.Operation steps1. Add 400 µL of saliva sample or saliva/preservation solution mixture.Note: 1) Saliva mixtures added to the preservation solution require a 50°C water bath for 1 hour or an empty 50°C temperature chamber for 2 hours prior to extraction.2) If an increase in sample volume is required, multiply the volumes of Proteinase K, Buffer GL, and anhydrous ethanol in Steps 2-4, and the liquid can be transferred in multiple times in Step 5.2. Add 40 µL of Proteinase K.3. Add 400µL Buffer GL, vortex and shake to mix thoroughly, and water bath at 56℃ for 15-30 minutes.Note: If RNA removal is required, add 4 µL of RNase A solution at a concentration of 100 mg/mL after the above steps are completed, vortex for 15 seconds, and leave at room temperature for 2 minutes.4. Centrifuge briefly to remove water droplets from the inside of the tube cap. Add 400 µL of anhydrous ethanol and mix well by vortexing and shaking. Centrifuge briefly.Note: 1) Vortex and shake to mix immediately after adding Buffer GL and anhydrous ethanol.The addition of Buffer GL and anhydrous ethanol may produce a white precipitate that will not affect subsequent experiments.2) A sol-gel product may be formed after GL and anhydrous ethanol, in which case vigorous shaking or vortexing is recommended.3) The solution obtained in the previous step is added to the adsorption column in the Collection Tube.5. (Spin Column DM) in the collection tube, and if the solution cannot be added all at once, it can be transferred in several times. centrifuge at 12,000 rpm (∼13,400 × g) for 1 min, pour off the waste solution in the collection tube, and put the adsorption column back into the collection tube.6. Add 500 µL of Buffer GW1 to the adsorption column (check that anhydrous ethanol has been added before use), centrifuge at 12,000 rpm for 1 minute, pour off the waste liquid in the collection tube, and put the adsorption column back into the collection tube.7. Add 500 µL of Buffer GW2 to the adsorption column (check that anhydrous ethanol has been added before use), centrifuge at 12,000 rpm for 1 minute, pour off the waste liquid in the collection tube, and put the adsorption column back into the collection tube.Note: Step 7 can be repeated if further DNA purity is required.8. 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 subsequent enzymatic reactions (digestion, PCR, etc.).9. Place the adsorption column in a new centrifuge tube (supplied), add 50-200 µL of Buffer GE or sterilized water to the middle of the adsorption column overhanging the column, let it stand at room temperature for 2-5 minutes, and centrifuge at 12,000 rpm for 1 minute to collect the DNA solution.-20°C to preserve DNA.Note: 1) If the downstream experiment is sensitive to pH or EDTA, you can use sterilized water for elution. The pH of the eluent has a great influence on the elution efficiency, if water is used as the eluent should ensure that its pH is 7.0-8.5 (you can use NaOH to adjust the pH of the water to this range), and the elution efficiency is not high when the pH is lower than 7.0.2) Buffer GE preheated in a 65-70°C water bath and incubated at room temperature for 5 min before centrifugation can increase the yield.3) Because DNA preserved in water is subject to acidic hydrolysis, for long-term storage, elution with Buffer GE and storage at -20°C is recommended... Read More |