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Geno-Serum Hepatitis B Kit R For Hepatitis B DNA purification from Serum Geno-Serum Hepatitis B Kit R provides a simple and ready to use system for the isolation of Hepatitis B DNA from serum at ambient temperature. The technology based on a filed US patent (US 61/538,566) and some proprietary products. The Kit contains tubes that can do a sequential vial lysis, extraction, binding, and elution of the DNA from hepatitis B in a single tube. Unlike other method and kits, DNA purified by Geno-Serum Hepatitis B Kit doesn t contain any inhibitory molecules for down-stream procedures such as qpcr or PCR reaction. The whole procedure takes less than 20 min with two short centrifugations. The Kit requires just one pippeting for whole DNA preparation in a single tube, which minimizes unnecessary cross-contamination between samples. The purified DNA is ideal for diagnosis of Hepatitis B infection using a serum. Geno-Serum Hepatitis B Kit R 2
Application ž ž Hepatitis B DNA preparation from serum and plasma. Ideal for diagnosis of Hepatitis B infection Features 1. The whole DNA extraction in 20 min. 2. Applicable for 0.2 ml of less volume of serum or plasma 3. The purified DNA doesn t have any inhibitory molecule for qpcr or PCR 4. The Kit allows more than 50 sample DNA preparations in an hour even without any automatic system. 5. The Kit can be stored at ambient temperature for more than a year without lose of activity. 6. The whole extraction in a single tube with a minimum sample handling 7. Minimize unnecessary cross-contamination between samples 8. By determining CT value of internal control (such as Actin or Beta-globulin gene), the success of whole DNA preparation procedure can be monitored. 9. Contains no organic solvent such as phenol or chloroform Geno-Serum Hepatitis B Kit R 3
Simple protocol Geno-Serum Hepatitis B Kit R 4
Procedures Isolation of hepatitis B viral DNA All procedures can be done in ambient temperature Preparations before you start: ž Set a heating block to 80 ž Pulse spin the sample extraction tube before use 1. Add 0.2 ml of serum to supplied hepatitis B extraction tube 2. Vortex 10 sec. and incubate at 80 o C for 10 min 3. Add 1 ml of 100% isopropanol and spin at 13,000 rpm for 5 min 4. Discard the supernatant by tilting the tube. 5. Add 1.0 ml of 70% EtOH to the microtube and vortex well until even suspension 6. Centrifuge at 13,000 rpm for 2 min 7. Discard the supernatant by tilting the tube 8. Remove all supernatant by a quick spin and pipetting 9. Add 100 μlof water and dissolve well by tapping 10. Take 5 μl for qpcr or PCR reaction Remarks 1. The recovered nucleic acid may contain some salts that carried from the solution. Although the salt doesn t interfere with any down-stream procedures of RT-PCR, PCR, cloning, Northern, Southern blotting, and sequencing, additional two more washing with 70% EtOH will completely remove any residual salt in the sample. 2. The dissolved pellet might have some debris that doesn t interfere with any downstream procedures such as qpcr or PCR. Geno-Serum Hepatitis B Kit R 5
Standard condition and procedure for Real Time PCR A. qpcr condition (1) Prepare the following qpcr reaction mixture. <per rxn> Reagents Volume(μl) Final concentration SYBR PCR premixture (2x) 12.5 1x PCR Forward Primer (10 µm) 0.5 0.2 µm PCR Reverse Primer (10 µm) 0.5 0.2 µm Template DNA from Geno-Kit 5 dh 2 O 6.5 Total 25 (2) qpcr Standard Protocol (50 cycles) Step Temperature Time Detection Initial Denaturation 95 C 10 sec. Off Denaturation 95 C 5 sec. Off Annealing / Extension 60 C 20 sec. On B. Example of the test using 0.2 ml of serum * 1 The values represent the CT value in qpcr reaction. Lower value indicates more copy number of target DNA. Samples* 1 Extraction method CT value HBV* 2 HBV Serum 0.2 ml Geno-Hepatitis B Kit 25.93 Qiaamp Circulating Nucleic Acid Prep Kit 27.23 *1; serum from hepatitis B infected patient. Qiaamp DNA mini Kit 27.24 *2; homemade primers for detection of hepatitis B virus gene Geno-Serum Hepatitis B Kit R 6
Specification Geno-Serum Hepatitis B Kit R for 48 reactions Component Geno-Serum Hepatitis B Tube Manual Contents 48 tubes/2 boxes/1 set 1 ea Materials required Bench top microcentrifuge Heat block (setting to 80 o C) 70% ethanol Quality Control In accordance with Genolution s Quality Management System, each lot of Genolution s Geno-Serum Hepatitis B Kit R is tested against predetermined specifications to ensure consistent product quality. Product Warranty and Satisfaction Guarantee Genolution Pharmaceuticals, Inc., guarantees the performance of all products in the manner described in our product manual. The customer must determine the suitability of the product for its particular use. We reserve the right to change, alter, or modify any product to enhance its performance and design. Technical Assistance Genolution s Technical Service Department is staffed by experienced scientists with extensive practical and theoretical expertise in sample and assay technologies and the use of Genolution products. If you have any questions or experience any difficulties regarding Genolution s Geno-Serum Hepatitis B Kit R or Genolution products in general, please do not hesitate to contact us. Genolution customers are a valuable source of information regarding advanced or specialized uses of our products. This information is helpful to other scientists as well as to the researchers at Genolution. We therefore encourage you to contact us if you have any suggestions about product performance or new applications and techniques. For technical assistance and more information, please contact our Technical Support Team at (02-3010- 8670, 8672) or visit Genolution website (www.genolution1.com) or through email at genolution@gmail.com. Geno-Serum Hepatitis B Kit R 7
Geno-Serum Hepatitis B Kit R 8