| Description | The introduction of Poly A tail at the 3'end of mRNA by tailing enzyme can increase the stability of mRNA and increase the efficiency of mRNA translation. The advantage of using the tailing enzyme to introduce the Poly A tail is that it is simple and easy to implement, avoiding the instability of The introduction of Poly A tail at the 3'end of mRNA by tailing enzyme can increase the stability of mRNA and increase the efficiency of mRNA translation. The advantage of using the tailing enzyme to introduce the Poly A tail is that it is simple and easy to implement, avoiding the instability of the plasmid caused by the introduction of too many consecutive T bases during plasmid construction.E. coli Poly (A) Polymerase does not rely on the presence of a template, and can catalyze the incorporation of ATP into the 3´ end of RNA in the form of AMP, that is, a poly A tail is added to the 3´ end of RNA. Poly (A) polymerase has a high tailing efficiency and can add 20~200 A bases to the 3´ end of RNA. Poly (A) structure helps to improve the translation efficiency of mRNA.This product is expressed in large-scale fermentation using Escherichia coli, is produced with medicinal specifications of raw materials, and strictly controls host protein residues, nucleic acid residues, etc., and conforms to GMP standard product production and quality management procedures to ensure that the production process and all raw materials can be traced.Quality requirements Project Standard Method Exterior Clear liquid Visual inspection Visible foreign body Compliance Chinese Pharmacopoeia 2020 Edition Fourth Part 1 Lamp Inspection Method (General Rule 0904) pH value 7.0-8.0 Chinese Pharmacopoeia 2020 Edition Part IV pH Determination Method (General Principle 0631) Active 4.9KUml-5.1KU/ml Tailing modification and efficiency determination method Purity ≥95% Chinese Pharmacopoeia 2020 Edition Part IV High Performance Liquid Chromatography (General Principle 0512) Endonuclease residue 004-DNA degradation does not exceed 10% Incubate SU enzyme with 004-DNA.37C for 3h Exonuclease residue 019 HindⅢ-DNA degradation does not exceed 10% Incubate SU enzyme with 019 HindⅢ-DNA at 37°C for 3h RNase residue Degradation of 293-RNA does not exceed 10% 5U enzyme and 293-RNA. Incubate at 37°C for 1h Bacterial endotoxin content ≤10 EU/mg Chinese Pharmacopoeia 2020 Edition Fourth Gel Limit Test Method (General Rule 1143) Host DNA residue ≤100 pg/mg Fluorescence quantitative PCR Host protein residue ≤50 ppm Chinese Pharmacopoeia 2020 Edition Part IV Method for the Determination of Bacterial Protein Residues (General Rule 3412) Mycoplasma Feminine Mycoplasma detection kit Heavy metal residue ≤10 ppm Chinese Pharmacopoeia 2020 Edition Fourth Heavy Metal Inspection Method (General Principle 0821) Follow the following specifications for production1. ISO 9001:2015, certified facility.2. "GMP Appendix-Cell Therapy Products" State Drug Administration.3. "General Introduction to Human Gene Therapy-Chinese Pharmacopoeia 2020" National Pharmacopoeia Commission.4. USP Chapter <1043>, Ancillary Materials for Cell, Gene, and Tissue-Engineered Products are used as excipients in cell therapy, gene therapy and tissue engineering products.5. USP Chapter <92>, Growth Factors and Cytokines Used in Cell Therapy Manufacturing Cytokines and growth factors used in the production of cell therapy products.6.Ph. Eur. General Chapter 5.2.12, Raw Materials of Biological Origin for the Production of Cell-based and Gene Therapy Medicinal Products.Product Usage1. Used for RNA 3´ end labeling.2. Add Poly(A) tail to RNA for cloning or affinity purification. For example, adding Poly (A) to miRNA provides an oligo-dT primer binding site for cDNA synthesis.3. Improve the translation efficiency of RNA in eukaryotic cells.Preservation system20mM Tris-HCl; 300mM NaCl; 1mM DTT; 1mM EDTA; 0.1% TritonX-100; 50% (v/v) Glycerol, pH 7.5.ApplicationsThe intact mRNA expresses GFP protein in the cell, the capping enzyme is compared with the cap analogPrecautions1. Thermal inactivation conditions: 65°C, 20 min.2. The enzyme can only use RNA as a substrate.3. The enzyme adds AMP to the 3´ end of RNA with high selectivity, and does not add the same length of Poly (A) to all RNA molecules.4. The enzyme uses M-MuLV reverse transcriptase reaction buffer, which can also be used for reaction.5. The enzyme requires divalent cations such as Mg2+ to be active.6. The length of RNA plus A tail is affected by the amount of enzyme, ATP, reaction time and other factors. The amount of A required for different experiments will be different. The length of A can be adjusted by reducing the reaction time. The enzyme is at 37°C. About 30 A bases can be added in 30 minutes of reaction, and about 100 A bases can be added in 1 hour.7. EDTA inhibits the enzyme activity. If the reaction is stopped, it can be purified directly by adding EDTA to a final concentration of 10mM... Read More | Product contentG665787Component5 mLStorageG665787A2×GoldStar Probe Mixture (UNG)5×1 mL-20℃. Avoid freeze/thaw cycle.G665787B50×High ROX200 µL-20℃. Avoid freeze/thaw cycle.G665787CddH2O 5×1 mL -20℃. Avoid freeze/thaw cycle. Product Introduction2× Product contentG665787Component5 mLStorageG665787A2×GoldStar Probe Mixture (UNG)5×1 mL-20℃. Avoid freeze/thaw cycle.G665787B50×High ROX200 µL-20℃. Avoid freeze/thaw cycle.G665787CddH2O 5×1 mL -20℃. Avoid freeze/thaw cycle. Product Introduction2× GoldStar Probe Mixture (UNG) is a premixed system dedicated to real-time fluorescence quantitative PCR by probe method (TaqMan, Molecular Beacon, etc.), with a concentration of 2×, containing GoldStar Taq DNA polymerase, PCR Buffer, dNTPs (dTTP is all replaced by dUTP), UNG enzyme and Mg2+, which is easy and convenient to operate. It is mainly used for the detection of genomic DNA target sequences and cDNA target sequences after RNA reverse transcription, such as gene expression analysis, copy number analysis and SNP genotype analysis. This product utilizes the dUTP-UNG anti-pollution system, which adds dUTP during the preparation of the PCR reaction system, thus forming an amplification product containing dU bases. This product can be eliminated by the UNG enzyme in the PCR system before the next PCR reaction. This effectively removes residual contamination of the PCR product and greatly reduces false positives due to contamination of the amplification product.UNG enzyme can be inactivated at the pre-denaturation step in the PCR cycle, and therefore will not affect the formation of new PCR products containing dU bases. The GoldStar Taq DNA Polymerase contained in this product is a chemically modified, new high-efficiency hot-start enzyme, which has no polymerase activity at room temperature, effectively avoiding non-specific amplification due to non-specific binding of primers and templates or primer dimerization at room temperature, and the activation of the enzyme must be incubated at 95°C for 10 minutes. The unique combination of PCR buffer system and hot-start enzyme significantly improves the amplification efficiency of PCR with stronger fluorescent signal and higher sensitivity to detect single-copy templates. A wider linear range and more accurate quantification of the target gene can be obtained by using this product.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 (G670150):Roche LightCycler 480, Roche LightCyler 96, Bio-rad iCyler iQ, iQ5, CFX96 and others.Instruments requiring Low ROX calibration(G665780):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 requiring High ROX calibration(G665787):ABI Prism 7000/7300/7700/7900, Eppendorf, ABI Step One/Step One Plus, and others.matters needing attentionBefore use, please mix gently by turning up and down, avoid foaming as much as possible, and use after brief centrifugation.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×GoldStar Probe Mixture（UNG）25 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, better results can be obtained with a primer concentration of 0.2 µM, and 0.1-1.0 µM can be used as a reference for setting the range.(2) The concentration of the probe used is related to the fluorescence quantitative PCR instrument used, the type of probe, and the type of fluorescent labeling substance, please refer to the instrument manual or the specific requirements for 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 programCaution! The pre-denaturation reaction of this product must be completed at 95°C for 10 minutes!Two-step PCR:Note: 1) The hot-start enzyme used in this product must be activated under the condition of pre-denaturation 95℃, 10min. 2) It is recommended to use two-step PCR reaction program, if you can't get good experimental results due to the use of primers with lower Tm value, etc., you can try to carry out three-step PCR amplification.Three-step PCR:... Read More | Inquire | Usually used industrially for the resolution of chiral compounds and the transesterification production of biodiesel | Purity> 96% by SDS-PAGE and HPLC analyses.FunctionHas weak activities on human monocytes and acts via receptors that also recognize MIP-1 alpha. It induced intracellular Ca(2+) changes and enzyme release, but no chemotaxis, at concentrations of 100-1,000 nM, and was inactive on T-lymphocytes, Purity> 96% by SDS-PAGE and HPLC analyses.FunctionHas weak activities on human monocytes and acts via receptors that also recognize MIP-1 alpha. It induced intracellular Ca(2+) changes and enzyme release, but no chemotaxis, at concentrations of 100-1,000 nM, and was inactive on T-lymphocytes, neutrophils, and eosinophil leukocytes. Enhances the proliferation of CD34 myeloid progenitor cells. The processed form HCC-1(9-74) is a chemotactic factor that attracts monocytes eosinophils, and T-cells and is a ligand for CCR1, CCR3 and CCR5.Post-translationalThe N-terminal processed forms HCC-1(3-74), HCC-1(4-74) and HCC-1(9-74) are produced in small amounts by proteolytic cleavage after secretion in blood. HCC-1(1-74), but not HCC-1(3-74) and HCC-1(4-74), is partially O-glycosylated; the O-linked glycan consists of one Gal-GalNAc disaccharide, further modified by two N-acetylneuraminic acids... Read More |