磁珠法血液基因组提取试剂盒基于磁珠分离纯化方式，适合从血液样品中分离纯化高质量基因组 DNA，提取过程中无需使用有毒的酚氯仿抽提，也无需进行耗时的醇类沉淀。该试剂盒可整合磁棒法或移液法自动核酸提取仪进行高通量提取实验，也可使用磁力分离架进行手工操作。
试剂盒纯化系统以超顺磁性的纳米磁性粒子为基质，这种磁性粒子在高浓度离液剂的条件下可通过氢键和静电特异的吸附核酸，而蛋白质或其它非特异吸附的少量杂质经洗涤被去除，最后用低盐缓冲液或 RNase-Free ddH₂O 洗脱核酸。纯化的核酸可适用于各种常规操作，包括 PCR、荧光定量 PCR 和二代测序检测等各种下游实验。
储存条件
试剂盒中所有组分可在室温干燥条件下保存 15个月。
产品组分表：
| D1372302 | Component | 50 T | Storage | | D1372302A | Buffer KL | 35 ml | RT | | D1372302B | Buffer DW | 55 ml | RT | | D1372302C | 磁珠XT | 1.2 ml | RT | | D1372302D | Proteinase K | 1.2 ml | RT | | D1372302E | Buffer EB | 15 ml | RT |
|
注意事项
1. 样品应避免反复冻融，否则会导致核酸提取量降低。样本可立即进行提取，也可于 4°C 保存待测，保存时间不超过 24 小时。长期保存可置于-20°C 或-80°C 冷冻保存。
2. 本试剂盒提取 DNA，操作过程注意防止核酸降解。操作人员应戴无粉手套、口罩等。
3. 使用前请详细阅读使用说明，严格按照使用说明书操作，临床样本等需在超净台或生物安全柜中进行。
4. 妥善处置所用样本及试剂材料，彻底清洗并消毒所用操作台面。
手工操作步骤
1. 向 1.5 ml 离心管中，加入 20 μl Proteinase K，之后加入 200 μl 血液。
2. 向离心管中加入 20 μl 磁珠 XT 和 600 μl Buffer KL，涡旋振荡 5 sec 使其充分混匀后，将离心管放于 56℃ 水浴锅中孵育 15 min，期间涡旋振荡混匀 2 次。
3. 将离心管放置于磁力架上静置 3 min，待磁珠完全吸附后，彻底弃去溶液（保持离心管固定于磁力架上）。
4. 将离心管从磁力架上取下，加入 500 μl Buffer DW 后涡旋点震 1 min 或涡旋振荡 5 sec 后放入 25℃、1500 rpm 的恒温混匀仪上振荡混匀 2 min (振荡过程中确保磁珠处于混匀状态)。之后将离心管放于磁力架上静置 1 min，待磁珠完全吸附于离心管侧壁后轻轻颠倒磁力架将离心管盖上的杂质洗落后彻底弃去溶液 (保持离心管固定于磁力架上)。
5. 重复步骤 4。
6. 将离心管从磁力架上取下，加入 500 μl 75% 乙醇（客户自备）后涡旋点震 1 min 或涡旋振荡 5 sec 后放入 25℃、1500 rpm 的恒温混匀仪上振荡混匀 2 min（振荡过程中确保磁珠处于混匀状态）。之后将离心管放于磁力架上静置 1 min，待磁珠完全吸附于离心管侧壁后轻轻颠倒磁力架将离心管盖上的杂质洗落后彻底弃去溶液（保持离心管固定于磁力架上）。
7. 保持离心管固定于磁力架上，用移液器进一步去除离心管管底和管盖上的溶液，之后室温放置 5-10 min，使乙醇挥发干净。
8. 将离心管从磁力架上取下，加入 50-100 μl Buffer EB。涡旋振荡使磁珠完全悬浮于洗脱液中后将其放于 56℃、1500 rpm 的恒温混匀仪上振荡洗脱 10 min，或将离心管放于 56℃水浴锅中孵育 10 min，期间每隔 3 min 涡旋振荡 10 sec。
9. 将离心管放于磁力架上静置 2 min，待磁珠完全吸附于离心管侧壁后用移液器将洗脱液转移至新的离心管中 - 20℃保存备用。
The magnetic bead-based blood genomic DNA extraction kit utilizes a magnetic separation and purification approach, suitable for isolating and purifying high-quality genomic DNA from blood samples. The extraction process eliminates the need for toxic phenol-chloroform extraction, as well as time-consuming alcohol precipitation. This kit can be integrated with magnetic rod-based or pipette-based automated nucleic acid extractors for high-throughput extraction experiments, and manual operation using a magnetic separation rack is also supported.
The purification system uses superparamagnetic nano-sized magnetic particles as the matrix. Under high-concentration chaotropic conditions, these magnetic particles specifically adsorb nucleic acids through hydrogen bonding and electrostatic interactions, while proteins and other non-specifically adsorbed minor impurities are removed through washing. Nucleic acids are finally eluted with low-salt buffer or RNase-Free ddH₂O. The purified nucleic acids are compatible with a wide range of routine downstream applications, including PCR, quantitative PCR, and next-generation sequencing assays.
Storage Conditions
All components of this kit can be stored at room temperature under dry conditions for 15 months.
Kit Components
D1372302
| Component | 50T | Storage |
|---|
| D1372302A | Buffer KL | 35 mL | RT | | D1372302B | Buffer DW | 55 mL | RT | | D1372302C | Magnetic Beads XT
| 1.2 mL | RT | | D1372302D | Proteinase K | 1.2 mL | RT | | D1372302E | Buffer EB | 15 mL | RT |
|
Precautions
1. Samples should avoid repeated freeze-thaw cycles, as this will lead to reduced nucleic acid yield. Samples can be processed immediately or stored at 4°C for up to 24 hours before testing. For long-term storage, keep at -20°C or -80°C.
2. During DNA extraction with this kit, precautions should be taken to prevent nucleic acid degradation. Operators should wear powder-free gloves and a face mask.
3. Please read the instructions thoroughly before use and follow the protocol strictly. Clinical samples must be handled in a biosafety cabinet or laminar flow hood.
4. Dispose of all samples and reagent materials properly, and thoroughly clean and decontaminate the work surface after use.
Manual Protocol
1. Add 20 μL Proteinase K to a 1.5 mL centrifuge tube, followed by 200 μL blood sample.
2. Add 20 μL Magnetic Beads XT and 600 μL Buffer KL to the tube. Vortex for 5 sec to mix thoroughly, then incubate at 56°C in a water bath for 15 min, with vortex mixing twice during incubation.
3. Place the tube on a magnetic rack and let stand for 3 min. Once the beads are fully separated, completely discard the supernatant (keep the tube fixed on the magnetic rack).
4. Remove the tube from the magnetic rack, add 500 μL Buffer DW, and either pulse-vortex for 1 min or vortex for 5 sec, then place in a thermomixer at 25°C, 1500 rpm for 2 min of shaking (ensure beads remain suspended during shaking). Then place the tube on the magnetic rack and let stand for 1 min. Once the beads are fully adsorbed to the side wall of the tube, gently invert the magnetic rack to wash off any impurities on the tube lid, then completely discard the supernatant (keep the tube fixed on the magnetic rack).
5. Repeat step 4.
6. Remove the tube from the magnetic rack, add 500 μL 75% ethanol (user-provided), and either pulse-vortex for 1 min or vortex for 5 sec, then place in a thermomixer at 25°C, 1500 rpm for 2 min of shaking (ensure beads remain suspended during shaking). Then place the tube on the magnetic rack and let stand for 1 min. Once the beads are fully adsorbed to the side wall of the tube, gently invert the magnetic rack to wash off any impurities on the tube lid, then completely discard the supernatant (keep the tube fixed on the magnetic rack).
7. Keep the tube fixed on the magnetic rack, use a pipette to further remove any residual solution from the bottom and lid of the tube, then leave at room temperature for 5–10 min to allow complete evaporation of residual ethanol.
8. Remove the tube from the magnetic rack, add 50–100 μL Buffer EB. Vortex to fully resuspend the beads in the elution buffer, then elute at 56°C in a thermomixer at 1500 rpm for 10 min. Alternatively, incubate at 56°C in a water bath for 10 min, with vortexing for 10 sec every 3 min.
9. Place the tube on the magnetic rack and let stand for 2 min. Once the beads are fully adsorbed to the side wall of the tube, transfer the eluate to a new centrifuge tube using a pipette and store at -20°C for later use.
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