计算溶液所需的质量、体积或浓度。
琼脂糖凝胶DNA回收试剂盒
有货
为什么选择此级别
BioReagent,分子生物学级 ，适用于对基线干扰要求严格的色谱和分析工作流程。
储存与运输
室温。常规运输 。请查阅批次 COA 获取详细规格。
质量文档
SDS、COA、产品数据表及规格说明书均可下载。可通过批号查询获取批次 COA。
文献证明
在色谱分析、有机合成和交叉偶联反应领域已被 0 篇同行评审文献引用。
概述
本试剂盒基于离心吸附柱技术，适合于从TAE或TBE琼脂糖凝胶上回收DNA片段，同时除去蛋白质、其它有机化合物、无机盐离子及寡核苷酸引物等杂质。使用本试剂盒可回收 100 bp-15 kb 大小的片段，回收率可高达80%，每个离心吸附柱每次可吸附的DNA量为10 μg，纯化的DNA可直接用于测序、连接、酶切、PCR和转化等。
操作步骤
操作步骤
使用前请先在Buffer PW2中加入一定量的无水乙醇。12 mL的Buffer PW2中加入48 mL无水乙醇，24 mL的Buffer PW2中加入96 mL无水乙醇。
1. (可选) 柱平衡步骤：向吸附柱 KOS 中 (吸附柱放入收集管中) 加入 500 μL 的 Buffer BL，12,000 rpm (13,680 × g) 离心 30 sec，弃废液，将吸附柱放回收集管中。(请使用当天处理过的吸附柱)
注意：长期室温存放的吸附柱经 Buffer BL 处理可以增加结合产量。
2. 将单一的目的DNA条带从琼脂糖凝胶中切下（尽量切除多余部分）放入干净的离心管中，称取重量。
3. 向胶块中加入3倍体积Buffer PBD（如果凝胶重为0.1 g，其体积可视为100 μL，则加入300 μL Buffer PBD），50℃溶胶5-10 min左右，其间不断温和地上下翻转离心管，以确保胶块充分溶解。（若胶块的体积过大，可事先将胶块切成碎块）。
注意：胶块完全溶解后最好将溶液温度降至室温再上柱，因为吸附柱在室温时结合DNA的能力较强。
4. 将上一步所得溶液加入到吸附柱KOS中（吸附柱放入收集管中），室温放置2 min，12,000 rpm (13,680 × g) 离心30 sec，弃废液，将吸附柱放回收集管中。
注意：吸附柱容积为700 μL，若样品体积大于700 μL可分次加入。
5. 向吸附柱中加入580 μL Buffer PW2（使用前请先加入无水乙醇），12,000 rpm (13,680 × g) 离 心30 sec，弃废液，将吸附柱放回收集管中。
注意：如果回收的DNA是用于盐敏感的实验，例如平末端连接实验或直接测序，建议PW2加入后静置2min再离心。
6. 重复步骤5。
7. 12,000rpm (13,680 × g) 离心2 min。
8. 将吸附柱放入一个干净离心管中，室温开盖晾干2 min，以彻底挥发乙醇。向吸附膜的中间部位悬空滴加适量的Buffer EB，12,000 rpm (13,680×g) 离心30 sec，收集DNA溶液。
注意：洗脱液的体积不应小于30 μL，体积过小会影响回收的效率。如果下游实验对pH值敏感，可以用灭菌水洗脱。洗脱液的pH值对洗脱效率有很大影响，若用水做洗脱液应保证其pH值在7.0 - 8.5（可以用NaOH将水的pH值调到此范围），pH值低于7.0时洗脱效率不高。为了提高DNA的回收量，可将离心得到的洗脱液重新加回离心吸附柱中，12,000 rpm (13,680×g) 离心30 sec，将DNA溶液收集到离心管中。
DNA浓度及纯度检测
纯化得到的DNA片段可用琼脂糖凝胶电泳和紫外分光光度计检测浓度与纯度。
通过测量分光光度计中260 nm处的吸光度（A₂₆₀）确定DNA浓度。如果需要稀释样本，建议用Buffer EB进行稀释，用水稀释样本可能导致数值不准确。260 nm处准确的吸光度应在 0.1-1.0 范围内。260 nm处1个单位的吸光度相当于50 µg/mL 双链DNA；相当于40 µg/mL 单链DNA。
A₂₆₀/A₂₈₀比值在 1.7-1.9 之间。如果洗脱时不使用Buffer EB，而使用ddH₂O，比值会偏低，因为pH值和离子存在会影响光吸收值，但并不表示纯度低。
This kit is based on centrifugal spin column technology and is suitable for recovering DNA fragments from TAE or TBE agarose gels while removing impurities such as proteins, other organic compounds, inorganic salt ions, and oligonucleotide primers. Using this kit, fragments ranging from 100 bp to 15 kb can be recovered with a recovery rate of up to 80%. Each spin column can bind up to 10 μg of DNA. The purified DNA can be directly used for sequencing, ligation, restriction enzyme digestion, PCR, and transformation.
Protocol
Before use, add an appropriate amount of absolute ethanol to Buffer PW2. For 12 mL of Buffer PW2, add 48 mL of absolute ethanol; for 24 mL of Buffer PW2, add 96 mL of absolute ethanol.
1. (Optional) Column Equilibration
Place the KOS spin column into a collection tube. Add 500 μL of Buffer BL to the spin column, centrifuge at 12,000 rpm (13,680 × g) for 30 seconds. Discard the waste liquid and put the spin column back into the collection tube. (Use spin columns processed on the same day.)
Note: Treating spin columns stored at room temperature for a long time with Buffer BL can improve binding yield.
Cut the single target DNA band from the agarose gel (remove as much extra gel as possible) and place it into a clean centrifuge tube. Weigh the gel slice.
Add 3 volumes of Buffer PBD to the gel slice (if the gel weighs 0.1 g, its volume can be considered as 100 μL, then add 300 μL of Buffer PBD). Dissolve the gel at 50°C for 5–10 minutes, gently inverting the tube occasionally to ensure complete dissolution. (If the gel volume is too large, cut it into small pieces beforehand.)
Note: After the gel is completely dissolved, it is best to cool the solution to room temperature before loading onto the column, as the spin column binds DNA more efficiently at room temperature.
Transfer the solution from the previous step to a Spin Column KOS (place the column in a collection tube). Let stand at room temperature for 2 minutes, then centrifuge at 12,000 rpm (13,680 × g) for 30 seconds. Discard the flow-through and place the column back into the collection tube.
Note: The column has a capacity of 700 μL. If the sample volume exceeds 700 μL, load in multiple batches.
Add 580 μL of Buffer PW2 (ensure absolute ethanol has been added before use) to the column. Centrifuge at 12,000 rpm (13,680 × g) for 30 seconds. Discard the flow-through and place the column back into the collection tube.
Note: If the recovered DNA is intended for salt‑sensitive experiments (e.g., blunt‑end ligation or direct sequencing), it is recommended to let the column stand for 2 minutes after adding PW2 before centrifugation.
6. Repeat step 5.
Centrifuge at 12,000 rpm (13,680 × g) for 2 minutes.
Place the column into a clean centrifuge tube and air‑dry it with the lid open at room temperature for 2 minutes to completely evaporate any residual ethanol. Add an appropriate volume of Buffer EB to the center of the membrane (without touching it), let stand for 2 minutes at room temperature, then centrifuge at 12,000 rpm (13,680 × g) for 30 seconds to collect the DNA solution.
Note: The elution volume should not be less than 30 μL, as smaller volumes may reduce recovery efficiency. If the downstream experiment is pH‑sensitive, sterile water can be used for elution. The pH of the elution buffer greatly affects elution efficiency; if using water, ensure its pH is between 7.0 and 8.5 (adjust with NaOH if necessary). A pH below 7.0 results in poor elution. To increase DNA yield, reload the eluate back onto the column and centrifuge again at 12,000 rpm (13,680 × g) for 30 seconds, collecting the DNA solution in the tube.
DNA Concentration and Purity Assessment
The purified DNA fragments can be analyzed by agarose gel electrophoresis and UV spectrophotometry to determine concentration and purity.
DNA concentration is determined by measuring the absorbance at 260 nm (A₂₆₀). If dilution is required, it is recommended to use Buffer EB; diluting with water may lead to inaccurate values. The accurate absorbance at 260 nm should be in the range of 0.1–1.0. An absorbance of 1 unit at 260 nm corresponds to 50 μg/mL for double‑stranded DNA and 40 μg/mL for single‑stranded DNA.
The A₂₆₀/A₂₈₀ ratio should be between 1.7 and 1.9. If ddH₂O is used instead of Buffer EB for elution, the ratio may be lower due to the influence of pH and ions on absorbance, but this does not indicate low purity.
规格
别名
DNA凝胶回收试剂盒 | DNA凝胶回收试剂盒（离心柱型）
英文别名
Gel Extraction Kit
规格或纯度
BioReagent, 分子生物学级
英文名称
DNA Gel Extraction Kit
应用
DNA Extraction
储存与运输
储存条件
室温
运输条件
常规运输
稳定性与储存
Store at room temperature long term (15 months).
组分与储存
| A1521384 | Component | 50T | 100T | Storage |
| A1521384A | Buffer PBD | 50 mL | 100 mL | RT. |
| A1521384B | Buffer PW2 | 12 mL | 24 mL | RT. |
| A1521384C | Buffer EB | 15 mL | 15 mL | RT. |
| A1521384D | 吸附柱KOS | 1 EA×50 | 1 EA×100 | RT. |
| A1521384E | 2 mL收集管 | 1 EA×50 | 1 EA×100 | RT. |
A1521384F | BL buffer | 30 mL | 60 mL | RT. |
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高级数据
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| 批号(Lot Number) | 证书类型 | 货号 |
|---|---|---|
 ZJ26F0636348 |
分析证书 | A1521384 |
技术文档和文章
溶液计算器
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