| Description | This product is a premixed system composed of Pfu DNA Polymerase, Mg2+, dNTPs, PCR stabilizers and enhancers, with a concentration of 2 ×. Pfu DNA Polymerase exhibits 5 ′ -3 ′ DNA polymerase activity and 3 ′ -5 ′ exonuclease activity, thus possessing error correction This product is a premixed system composed of Pfu DNA Polymerase, Mg2+, dNTPs, PCR stabilizers and enhancers, with a concentration of 2 ×. Pfu DNA Polymerase exhibits 5 ′ -3 ′ DNA polymerase activity and 3 ′ -5 ′ exonuclease activity, thus possessing error correction ability during DNA amplification. Compared with Taq DNA Polymerase, it has high fidelity (6-8 times that of Taq enzyme) and better thermal stability. The pre prepared PCR mixture makes the operation simpler and faster, minimizing human error and contamination to the greatest extent possible. The original MasterMix formula makes the entire reaction system very stable and has good repeatability. This product has been added with a dye (blue), and can be directly subjected to electrophoresis detection after the reaction is completed. The Pfu DNA polymerase contained in this product has the characteristics of low mismatch rate and high temperature resistance, making it suitable for gene cloning, gene directed mutagenesis, SNP and end effector complement reactions. P665594 Component 1 mL 5 mL Storage P665594A 2×Pfu MasterMix (Dye) 1 mL 5×1 mL -20℃. Avoid freeze/thaw cycle. P665594B ddH2O 1 mL 5×1 mL -20℃. Avoid freeze/thaw cycle. Notes: 2×Pfu MasterMix contains Pfu DNA Polymerase, 3mM MgCl2 and 400µM each dNTP. Quality control:After testing, there was no exogenous nuclease activity; PCR method for detecting residual DNA without host; Can effectively amplify single copy genes from multiple genomes; Storage at 2-8 ℃ for three months showed no significant change in activity.Usage:The following is an example of a PCR reaction system and reaction conditions for amplifying a 1 kb fragment using human genomic DNA as a template. In practical operation, corresponding improvements and optimizations should be made based on the template, primer structure, and target fragment size.1. PCR reaction system Reagent 50 µlReaction system Final concentration 2×Taq MasterMix（Dye） 25 µL 1× Forward Primer，10 µM 2 µL 0.4 µM Reverse Primer，10 µM 2 µl 0.4 µM Template DNA <0.5 µg <0.5 µg/50 µL ddH2O up to 50 µL /Attention: When amplifying with Pfu enzyme, the purity of the primers is required to be high, and the length of the primers is greater than 18 bases. The primer concentration should be based on the final concentration of 0.1-1.0 µ M serves as a reference for setting the range. In the case of low amplification efficiency, the concentration of primers can be increased; When non-specific reactions occur, the primer concentration can be reduced to optimize the reaction system.2. PCR reaction conditions Step Temperature Time / Pre denaturation 94℃ 2 min / Denaturation 94℃ 30 s 25-35 cycles Anneal 55-65℃ 30 s 25-35 cycles Extend 72℃ 60 s 25-35 cycles Finally extended 72℃ 5 min / Attention:1) The thermal stability of Pfu enzyme is better than that of Taq enzyme. For templates with high GC content, the denaturation temperature can be increased to 98 ℃ without affecting the activity of Pfu enzyme.2) In general experiments, if the annealing temperature is 5 ℃ lower than the melting temperature Tm of the amplification primer, and the ideal amplification efficiency cannot be achieved, the annealing temperature should be appropriately reduced; When non-specific reactions occur, increase the annealing temperature to optimize the reaction conditions.3) Pfu enzyme has 3 '-5' exonuclease activity, so the extension rate of Pfu enzyme during amplification is much lower than that of Taq enzyme. The extension time is set according to the size of the amplified fragment, and the amplification extension rate of this product is 1 kb/min.4) The number of cycles can be set based on the downstream application of the amplification product. If the number of cycles is too small, the amplification amount is insufficient; If there are too many cycles, the probability of mismatches will increase, and non-specific backgrounds will be severe. So, while ensuring product yield, the number of cycles should be minimized as much as possible.5) This product has 3 '-5' exonuclease activity. The PCR product is a flat end and cannot be directly used for T/A cloning. If T/A cloning is required, "A" needs to be added at its end or cloned using a flat end vector... Read More | Inquire | Store at +4°C. Store under desiccating conditions. The product can be stored for up to 12 months | Purity:>95%, by SDS-PAGE visualized with Coomassie® Blue Staining.Description：HSPD1, also known as HSP60, is a member of the chaperonin family. HSPD1 may function as a signaling molecule in the innate immune system. This protein is essential for the folding and assembly of newly Purity:>95%, by SDS-PAGE visualized with Coomassie® Blue Staining.Description：HSPD1, also known as HSP60, is a member of the chaperonin family. HSPD1 may function as a signaling molecule in the innate immune system. This protein is essential for the folding and assembly of newly imported proteins in the mitochondria. It may also prevent misfolding and promote the refolding and proper assembly of unfolded polypeptides generated under stress conditions in the mitochondrial matrix. HSPD1 gene is adjacent to a related family member and the region between the 2 genes functions as a bidirectional promoter. Several pseudogenes have been associated with this gene. Mutations associated with this gene cause autosomal recessive spastic paraplegia 13. Defects in HSPD1 are a cause of spastic paraplegia autosomal dominant type 13 (SPG13). Spastic paraplegia is a degenerative spinal cord disorder characterized by a slow, gradual, progressive weakness and spasticity of the lower limbs. Defects in HSPD1 are the cause of leukodystrophy hypomyelinating type 4 (HLD4); also called mitochondrial HSP60 chaperonopathy or MitCHAP-60 disease. HLD4 is a severe autosomal recessive hypomyelinating leukodystrophy. HSPD1 is clinically characterized by infantile-onset rotary nystagmus, progressive spastic paraplegia, neurologic regression, motor impairment, profound mental retardation. Death usually occurs within the first two decades of life... Read More | Product Characteristics UNI-StabilPLUS is a universal stabilizer for the dilution and stabilization of both Horseradish Peroxidase (HRP) and Alkaline Phosphatase (AP) labeled proteins and antibodies, in order to maintain the molecular conformation and prevent loss of activity over time. This enablesProduct Characteristics UNI-StabilPLUS is a universal stabilizer for the dilution and stabilization of both Horseradish Peroxidase (HRP) and Alkaline Phosphatase (AP) labeled proteins and antibodies, in order to maintain the molecular conformation and prevent loss of activity over time. This enables the making of pre-diluted, ready-to-use conjugates, minimizing assay errors in dilution. Superior stabilization of HRP and AP conjugated antibodies in low as well as high protein dilutions is seen, when using UNI-StabilPLUS. When tested with AP conjugated antibody stability is seen as follows: • at least 3 years at 2-8 °C • at least 2 years at room temperature • at least 4 weeks at 37 °C When tested with HRP conjugated antibody stability is seen as follows: • at least 2 years at 2-8 °C • at least 1 years at room temperature • at least 2 weeks at 37 °CUNI-StabilPLUS is recommended for the dilution of antibodies directed against rabbit immunoglobulins unlike HRP-StabilPLUS (cat. no. H494387) and Antibody Enhancer (cat. no. A494276).Composition & Properties UNI-StabilPLUS is a ready-to use buffer that appears as an opaque solution. The product is based on a mild acid Tris buffer containing proprietary stabilizing components. UNI-StabilPLUS contains neither BSA, nor other material from bovine serum, no azide, mercury or other toxic components.Working Procedure 1.Make a series of dilutions of the HRP- or AP conjugated protein in UNI-StabilPLUS in order to determine the optimal dilution. 2.Run the assay as usual or store the diluted conjugated protein preferably at 2-8 °C.Tips & Tricks • Avoid using phosphate buffers for AP-conjugated antibody assays. We recommend the use of Tris/HCl, Tween as the washing buffer, instead of a PBS buffer which will reduce signal significantly. • For extended stability of HRP conjugated antibodies, HRP-StabilPLUS (cat. no. H494387) is recommended. Handling & Storage • Store solution at 2-8 °C... Read More |