KAPA HiFi HotStart ReadyMix降低扩增偏差，提高测序覆盖度。

使用产品K（左）或产品C（右），从100 ng Covaris打断的人类基因组DNA构建文库用作全基因组鸟枪法测序。使用K的扩增偏差更低，覆盖更均匀。数据由The Broad Institute提供 (Cambridge, MA, USA)。

焦磷酸测序确定了KAPA HiFi酶行业领先的高保真度。

Taq酶、基于Taq酶的长片段PCR预混液、具有校正功能的DNA聚合酶(产品N和产品T)的已发表错误率，与KAPA HiFi HotStart DNA聚合酶的错误率比较结果(每3.54×10-6个核苷酸掺入1个错误，由454平台深度测序验证)。相比而言，KAPA HiFi酶比Taq酶低50-100倍。

KAPA HiFi Uracil+ReadyMix不受dUTP抑制。

SYBR® Green法实时荧光定量PCR检测dUTP对PCR扩增的影响，结果显示dUTP可以抑制常规PCR反应液的扩增(左)，而KAPA HiFi Uracil+ ReadyMix (右) 无扩增抑制(dUTP浓度为0.2 mM，扩增片段长度为452 bp，模板稀释比例为80 pM-0.8 fM)。

KAPA HiFi Uracil+ ReadyMix 提供更高的文库产量和最小的文库分子大小偏差。

使用12、14或16个循环的标准方案扩增人类全基因组亚硫酸氢盐处理的文库，并使用Bioanalyzer® 2100高灵敏度DNA检测系统分析扩增文库。与A相比，KAPA HiFi Uracil+ ReadyMix的产量更高 (左)，片段大小偏差更小 (右)。

产品手册

Kits can be stored for up to 12 months at -20˚C.

Kits include KAPA HiFi HotStart ReadyMix (2X), a convenient PCR master mix containing KAPA HiFi HotStart DNA Polymerase, KAPA dNTPs, reaction buffer, and MgCl2 at a final concentration of 2.5 mM. Kits with primers are also available.

Kits for real-time library amplification include a real-time version of the KAPA HiFi HotStart ReadyMix, as well as fluorescent standards to monitor library amplification.

构成要素

规格明细

KAPA文库定量试剂盒的工作原理。

qPCR的引物，根据平台特异性的文库街头（adaper)序列而设计，同时扩增6个预稀释的、组成浓度梯度的DNA标准品，以及稀释至合适浓度的待定量的NGS文库。将每个已知浓度DNA标准品的平均Cq值作图，生成标准曲线。标准曲线用于将稀释的待测NGS文库的平均Cq值转换为浓度，从而计算出待测文库的工作浓度。

准确、可重复的文库定量能使成簇密度达到高度一致。

在Broad研究所(Cambridge, MA, USA)，在Illumina® GA测序工作流程中应用KAPA文库定量试剂盒显著降低了簇密度的差异性，并消除了滴定的需求。此处展示了连续文库每个tile中簇的平均数目。

产品手册

Kits can be stored for up to 12 months at -20˚C.

Complete kits include KAPA SYBR FAST qPCR Master Mix (2X), Primer Premix (10X) and a set of 6 DNA Standards. Reagents for qPCR (KAPA SYBR FAST qPCR Master Mix and Primer Premix) and DNA Standards (1–6) are also sold separately. Where noted, Master Mixes contain instrument-specific reference dyes, while the Universal kit includes ROX High and ROX Low (both 50X) separately.

Components–Illumina® Platform

Components–Illumina® Platform

规格明细

高灵敏性的检测表达基因。

使用KAPA Stranded mRNA-Seq制备的文库，与 I 系列产品制备同类型文库相比有着更低的重复率，保持了链特异性的同时，更多特异的转录本和特异的基因被检出。

覆盖均一性提升。

500 ng高质量RNA使用KAPA Stranded mRNA-Seq Kit和 I 系列产品进行文库制备，KAPA文库制备试剂盒在5’和3 端’位置测序的均一性更好。

富含GC转录本的覆盖度得到提升。

DVL3转录本的5’和3’端位置外显子区域(红色)为高GC含量区域。使用KAPA StrandedmRNA-Seq试剂盒(绿色)测序结果与 I 系列产品(橙色)结果相比，绿色区域有着更高的测序覆盖度。

低表达的转录本的覆盖度提升。

使用KAPA Stranded mRNA-Seq试剂盒(绿色)与 I 系列产品(橙色)进行文库制备，KAPA在GLTPD1基因上的覆盖度更高，即使测序深度相同， I 系列产品覆盖度仍有缺失(红色区域)。

产品手册

Shipping and storage

The enzymes provided in this kit are temperature sensitive, and appropriate care should be taken during shipping and storage. KAPA Stranded RNA-Seq Library Preparation Kits are shipped on dry ice or ice packs, depending on the destination country. Upon receipt, immediately store enzymes and reaction buffers at -20˚C in a constanttemperature freezer. The PEG/NaCl SPRI^® Solution should be protected from light, and stored at -20˚C. For shortterm use, the PEG/NaCl SPRI^® Solution may be stored protected from light at 4˚C for up to 2 months. When stored under these conditions and handled correctly, the kit components will retain full activity until the expiry date indicated on the kit label.

Handling

Always ensure that components have been fully thawed and thoroughly mixed before use. Keep all reaction components and master mixes on ice whenever possible during handling. KAPA HiFi HotStart ReadyMix (2X) may not freeze completely, even when stored at -20 °C. Nevertheless, always ensure that the KAPA HiFi HotStart ReadyMix is fully thawed and thoroughly mixed before use. PEG/NaCl SPRI^® Solution does not freeze at -20 °C, but should be equilibrated to room temperature and mixed thoroughly before use.

Quality control

All kit components are subjected to stringent functional quality control, are free of detectable contaminating exoand endonuclease activities, and meet strict requirements with respect to DNA contamination. Please contact support@kapabiosystems.com for more information.