QIAquick PCR Purification Kit for PCR Cleanup

用于纯化小于 10 µg 的 PCR 产物（100 bp 至 10 kb）

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QIAquick PCR Purification Kit (50)

Cat. No. / ID: 28104

适用于 50 次 PCR 反应的纯化：50 个 QIAquick Spin Columns、缓冲液、Collection Tubes (2 ml)

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Kit离心柱

QIAquick PCR Purification Kit

QIAquick PCR & Gel Cleanup Kit

QIAquick Spin Columns

制备

250

QIAquick PCR Purification Kit 旨在用于分子生物学应用。该产品不能用于疾病诊断、预防和治疗。

✓ 全天候自动处理在线订单

✓ 博学专业的产品和技术支持

✓ 快速可靠的（再）订购

特点

即用型 DNA 的回收率高达 95%
只需三个简单步骤，即可净化长达 10 kb 的 DNA
凝胶加载染料方便样本分析
用于凝胶提取和 PCR 净化的组合试剂盒
可单独购买离心柱

产品详情

QIAquick PCR Purification Kit 提供离心柱、缓冲液和采样管，用于以硅胶膜为基础纯化大于 100 bp 的 PCR 产物。使用简单快速的结合-洗涤-洗脱程序和 30–50 μl 的洗脱体积，可纯化不高于 10 kb 的 DNA。可选的 pH 指示剂可轻松确定 DNA 吸附到离心柱的最佳 pH 值。程序可在 QIAcube Connect 上全自动完成。

为获得最佳效果，建议将本产品与 QIAvac 24 Plus 一起使用。

QIAquick PCR Purification 标准方案也可使用 TRACKMAN Connected 系统并搭配 PIPETMAN M Connected 移液管（二者均来自 Gilson）执行。TRACKMAN Connected 系统可指导研究人员完成 QIAquick PCR Purification 方案，同时自动调整支持蓝牙功能的 PIPETMAN M Connected 移液器设置。会记录实验方案执行的每一步骤，通过生成全面的运行报告以加快报告。下载更多信息.

绩效

QIAquick PCR 纯化程序可去除 DNA 样本中的引物、核苷酸、酶、矿物油、盐和其他杂质（见图“”）。使用微型离心机或真空歧管，可纯化出 100 bp 至 10 kb 的 DNA。

查看图表

PCR 后完全去除引物。

原理

QIAquick Kit 包含一个硅胶膜组件，用于在高盐缓冲液中结合 DNA，然后用低盐缓冲液或水进行洗脱。纯化程序可去除 DNA 样本中的引物、核苷酸、酶、矿物油、盐、琼脂糖、溴化乙锭和其他杂质（见图“pcr PCR 后完全去除引物”）。硅胶膜技术消除了树脂松散和稀浆带来的问题和不便。专用结合缓冲液针对特定应用进行了优化，可促进特定大小范围内 DNA 分子的选择性吸附。

凝胶加载染料

为了更快、更方便地进行样本处理和分析，我们提供了凝胶加载染料。GelPilot Loading Dye 包含三种跟踪染料（二甲苯胍青、溴酚蓝和橙黄 G），便于优化琼脂糖凝胶的运行时间，防止较小的 DNA 片段迁移过远（见图“ GelPilot Loading Dye”）。

查看图表

PCR 后完全去除引物。

GelPilot Loading Dye。

程序

QIAquick 系统使用简单的结合-洗涤-洗脱程序（见流程图“ QIAquick 和 MinElute 程序”）。将结合缓冲液直接加到 PCR 样本或其他酶促反应中，然后将混合物加到 QIAquick 离心柱上。结合缓冲液中含有 pH 指示剂，便于确定 DNA 结合的最佳 pH 值（见图 “pH 指示基团染料”）。核酸在缓冲液提供的高盐条件下吸附到硅胶膜上。杂质被洗去，用少量低盐缓冲液或水洗脱出的纯 DNA 可直接用于所有后续应用。

处理

QIAquick 离心柱的设计提供了两种方便的处理选项。离心柱可装入传统的台式微型离心机或任何带有鲁尔接头的真空歧管，如带有 QIAvac Luer Adapter 的 QIAvac 24 Plus。QIAquick PCR Purification Kit 以及其他基于 QIAGEN 离心柱的试剂盒可在 QIAcube 上实现全自动操作，从而提高生产效率和结果的规范化（见图“离心柱处理选项 A、 B、 C、 D”和“ QIAcube Connect”）。

查看图表

QIAquick 和 MinElute 程序。

应用

用 QIAquick 系统纯化的 DNA 片段可直接用于所有应用，包括测序、微阵列分析、连接和转化、限制性酶切、标记、微量注射、PCR 和体外转录。

辅助数据和图表

QIAquick 和 MinElute 程序。

QIAquick 和 MinElute 系统使用离心柱或真空歧管进行简单的结合-洗涤-洗脱程序。

Specifications

Features	Specifications
Binding capacity	10 µg
Technology	硅胶膜技术
Sample type: applications	来自 PCR 和其他酶促反应的 ssDNA 或 dsDNA
Fragments removed	< 40mer
Elution volume	> 30 µl
Fragment size	100 bp – 10 kb
Processing	手动
Format	试管
Type(s) of DNA recovered	ss DNA 和 dsDNA

资源

The protocol has been used successfully for Cy3-, Cy5-, and biotin-labeling of cDNA from <50 ng of total RNA or poly A⁺ mRNA.

Download Safety Data Sheets for QIAGEN product components.

FAQ

Larger DNA fragments bind more tightly to the QIAquick columns. It is difficult to predict whether a DNA fragment larger than 10 kb can be efficiently recovered, because this depends on base composition as well as fragment size. If the fragment is only a few kb larger than the 10 kb limit, it can be helpful to heat the elution buffer EB to 60°C and let it incubate on the column for a few minutes before centrifuging. However, please note that it will become less likely to recover your sample the larger the fragment size is. As we cannot guarantee recovery of fragments larger than the maximum cutoff size, we do not recommend to purify such fragments using QIAquick Kits.

The QIAEX II Kit can be used to extract DNA fragments up to 50 kb from agarose or polyacrylamide gels.

FAQ ID -756

Buffer PB of the QIAquick PCR Purification Kit cannot be used to extract DNA from agarose gels. However, Buffer QG of the QIAquick Gel Extraction Kit can be used to remove salt and proteins from enzymatic reactions by adding 3 volumes of Buffer QG and 1 volume of isopropanol to the reaction and proceeding with step 6 of the Gel Extraction Spin Protocol in the QIAquick Spin Handbook. See the QIAquick Spin Handbook for a list of reactions which can be cleaned up with the various QIAquick kits.

FAQ ID -786

No. While columns from the QIAprep Spin Miniprep Kit and the QIAquick PCR Purification- and Gel Extraction Kits are based on silica-gel-membrane technology, each is designed to work optimally within its own kit format. In addition, the binding capacity and DNA recovery size cut-offs of the QIAprep and QIAquick Spin columns are different. QIAprep Spin columns bind up to 20 ug of plasmid DNA up to 50 kb in length, while the QIAquick Spin columns bind up to 10 ug of DNA with a maximum fragment size of 10 kb.

FAQ ID -311

Yes - all QIAquick Spin Kits contain identical columns, but different binding buffers optimized for each specific application.

FAQ ID -577

Buffer PE did not contain ethanol

Ethanol must be added to Buffer PE (concentrate) before use. Repeat procedure with correctly prepared Buffer PE.

Inappropriate elution buffer

DNA will only be eluted efficiently in the presence of low-salt buffer (e.g., Buffer EB: 10 mM Tris·Cl, pH 8.5) or water. Elution efficiency is strongly dependent on the salt concentration and pH of the elution buffer. Contrary to adsorption, elution is most efficient under basic conditions and low salt concentrations. DNA is eluted with 50 or 30 µl of the provided Buffer EB (10 mM Tris·Cl, pH 8.5), or water. The maximum elution efficiency is achieved between pH 7.0 and 8.5. When using water to elute, make sure that the pH is within this range. In addition, DNA must be stored at –20°C when eluted with water since DNA may degrade in the absence of a buffering agent. Elution with TE (10 mM Tris·Cl, 1 mM EDTA, pH 8.0) is possible, but not recommended because EDTA may inhibit subsequent enzymatic reactions.

Elution buffer incorrectly dispensed

Add elution buffer to the center of the QIAquick membrane to ensure that the buffer completely covers the membrane. This is particularly important when using small elution volumes (30 µl).

FAQ ID -180

Although it is possible to use the QIAquick Nucleotide Removal Kit and QIAquick PCR Purification Kit for RNA purification, the conditions are not optimized for RNA, nor are RNase-free conditions guaranteed. The RNeasy MinElute Cleanup Kit is recommended for clean-up, concentration and desalting of RNAs above 200 bases in length.

FAQ ID -490

CoralLoad dyes supplied in PCR Kits such as, e.g., Taq, HotStarTaq, and TopTaq DNA Polymerase and TopTaq Master Mix do not interfere with most downstream enzymatic applications.

However, for reproducible results, purification of PCR products using the QIAquick or MinElute PCR Purification Kits prior to enzymatic manipulation is recommended.

FAQ ID -1745

Add 1/10 volume of 3 M Na-Acetate pH 5.2, and 2 to 2.5 volumes of ice-cold 100% ethanol to the DNA sample
Mix, and store at –20°C for at least 1 h to precipitate the DNA
Recover the precipitated DNA by centrifugation at full speed in a microcentrifuge for 15–20 min
Pour off the ethanol and wash the pellet twice with room-temperature 70% ethanol
Allow the DNA pellet to air-dry
resuspend the DNA in a suitable volume of sterile TE buffer or distilled water

FAQ ID -305

Yes, please follow the Supplementary Protocol 'Purification of DNA fragments from dye-labeled reactions using the QIAquick PCR Purification Kit' (QQ06).

FAQ ID -947

Buffer PB contains a high concentration of guanidine hydrochloride and isopropanol. The exact composition of Buffer PB is confidential. However, this buffer can be purchased separately: Buffer PB.

FAQ ID -2791

Based on our experience, the QIAquick PCR Purification Kit removes SYBR Green dye efficiently from PCR reactions.

We strongly assume that the QIAquick Gel Extraction Kit will be equally efficient in removing this dye, however, we recommend a 5 min incubation with wash Buffer PE on the QIAquick spin column at step 10 of the QIAquick Gel Extraction Kit Protocol. Efficiency of SYBR Green dye removal has to be validated by the enduser.

FAQ ID -637

No - mineral oil will not affect the clean-up procedure with the QIAquick or MinElute PCR Purification Kit.

FAQ ID -575

Yes. The QIAquick PCR Purification Kit has been used to clean up fragments between 100 bp and 10 kb from a wide range of enzymatic reactions, removing salts, buffers, enzymes, nucleotides, and primers smaller than 40 nucleotides. Reactions that can be cleaned up with the QIAquick PCR Purification Kit include restriction digests, random priming, ligase, kinase, phosphatase, nuclease, nick translation, and cDNA synthesis reactions.
For data and additional information, please see QIAGEN News article Issue No. 5, 1998 "Fast and efficient enzyme removal with QIAquick Spin kits."

FAQ ID -130

DNA fragments purified with the QIAGEN DNA Cleanup Systems, i.e., the QIAquick PCR Purification Kit, the MinElute Reaction Cleanup Kit, the QIAEX II Gel Extraction Kit etc. may float out of the loading wells of agarose gels due to residual ethanol carried over from the wash step with Buffer PE (despite the addtition of glycerol-containing loading buffer).

Use either of the following options to remove residual ethanol from the eluate:

re-purify the sample using a QIAquick-, or MinElute column, or QIAEX II resin
incubate the eluate at 56°C for 10 min to evaporate the ethanol
dry down the sample in a vacuum centrifuge, and resuspend the pellet in a small volume of sterile water

FAQ ID -205

Yes, bisulfite containing methylation reactions can be cleaned up with our silica-based cleanup products, such as QIAquick and QIAEX II. Please see Goyon et al. (1994), 'Perpetuation of cytosine methylation in Ascobolus immersus implies a novel type of maintenance methylase', published in J Mol Biol. 1994 Jul 1;240(1):42-51, for a reference.

FAQ ID -519

As a rule of thumb, single-stranded DNA binds to silica with approximately half the affinity of a double-stranded DNA fragment of the same length under the buffer conditions used in the QIAquick and MinElute Kits. Even though no systematic experimental data exists, we expect that recovery of ssDNA fragments of approximately 200 nucleotides and below will not be very efficient after cleanup using the QIAquick PCR Purification Kit or MinElute PCR Purification Kit. By comparison, it should be possible to purify fragments longer than 140 nucleotides using the QIAquick Gel Extraction Kit.

Note that recovery of single strand DNA is influenced to some degree also by factors such as base composition and secondary structure. It has to be determined empirically by the researcher if cleanup of single-stranded DNA with QIAquick columns yields satisfactory results.

FAQ ID -759

The composition of Buffer EB is:

10 mM Tris-Cl, pH 8.5

Buffer EB is the elution buffer used in the QIAquick PCR, Gel Extraction, Nucleotide Removal Kits, and MinElute Kits for DNA cleanup, and the QIAprep Miniprep Kits for small-scale plasmid purification. The purified DNA can also be eluted in TE (10 mM Tris-Cl, 1 mM EDTA, pH 8.0), but the EDTA may inhibit subsequent enzymatic reactions.

FAQ ID -199

Occasionally, DNA fragments eluted from the silica matrix of QIAquick, MinElute or QIAEX II Kits will contain denatured single-stranded DNA (ssDNA), appearing as a smaller band on an analytical gel. Under certain conditions, chaotropic agents (present in all silica-based DNA purification methods) can denature DNA fragments. This is a rare event that may be influenced by sequence characteristics such as the presence of inverted repeats or A–T-rich stretches.

Because salt and buffering agents promote renaturation of DNA strands, the following tips are recommended:

use the eluted DNA to prepare your downstream enzymatic reaction, but omit the enzyme. Incubate the reaction mix at 95°C for 2 minutes to reanneal the ssDNA, and allow the tube to cool slowly to room temperature before adding the enzyme and proceeding
alternatively, the DNA can be eluted from the silica-gel membrane or resin in 10 mM Tris buffer containing 10 mM NaCl. However, the salt concentration of the eluate must then be taken into consideration in downstream applications.

FAQ ID -148

The QIAquick Spin Columns (100) (cat. no. 28115) in the QIAquick PCR Purification, Gel Extraction, Nucleotide Removal and PCR & Gel Cleanup kits are also sold separately from the kits.

The MinElute columns in the MinElute PCR Purification, Gel Extraction and Reaction Cleanup kits are not sold separately.

We always provide extra buffers in our kits so you can scale up reactions, add extra washes or allow for spillage.

FAQ ID -2460