使用SYBR® Green进行两步法和一步法定量RT-PCR,用于基因表达分析

  • 适用于全基因组的预制引物确保结果可靠
  • PCR效率接近100%,可靠的定量检测
  • 灵敏度高、特异性好
  • 宽线性范围内的准确定量检测
  • SYBR Green检测方法可节约时间和成本
QuantiTect Primer Assays是生物信息学验证的全基因组引物对,可在任何PCR仪上进行基于SYBR Green的real-time RT-PCR。引物对覆盖所有人类、小鼠、大鼠和其他各类生物的基因。每个QuantiTect Primer Assay包含特定基因的正反引物对,以冻干粉形式提供,经重悬可获得10倍稀释的引物溶液(real-time RT-PCR反应液组分需单独订购)。QuantiTect Primer Assays配合QuantiFast、QuantiTect、Rotor-Gene或FastLane Kit进行SYBR Green检测,可在real-time RT-PCR中确保高特异性和灵敏度的结果,其表现与探针法检测相当。
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You can search for the following terms:
  • Entrez Gene IDs (e.g., 835)
  • RefSeq IDs (e.g., NM_032983, NP_116765)
  • Gene symbols (e.g., CASP2)
  • Cat. no. (e.g., SI00299551, QT01342509)
  • Sanger ID or Accession (e.g., hsa-let-7b, MI0000063)
  • CpG loci identification numbers (CG#) (e.g., CG17753661)

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QuantiTect Primer Assay (200)
For 200 x 50 µl reactions or 400 x 25 µl reactions: 10x QuantiTect Primer Assay (lyophilized) supplied in single tube
249900 Varies
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Reference assays for SYBR Green detection

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Mouse
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QuantiTect Primer Assays are intended for molecular biology applications. These products are not intended for the diagnosis, prevention, or treatment of a disease.


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Superior sensitivity in real-time RT-PCR.|Reproducible real-time RT-PCR.|Time savings in development of real-time RT-PCR assays.|
MAPK1 (a protein kinase) and IL6 (a cytokine) in human leukocyte cDNA (1 ng) were quantified in duplicate using [A] QuantiTect Primer Assays and the QuantiTect SYBR® Green PCR Kit, or [B] probe-based assays and a real-time PCR kit from Supplier AII. QuantiTect Primer Assays provided greater sensitivity than the probe-based assays, as demonstrated by the lower CT values, as well as high specificity, as indicated by the single peaks in melting curve analysis.|[A] Different amounts of leukocyte cDNA were analyzed in duplicate using the QuantiTect Primer Assay for IL8 (a chemokine) and QuantiFast SYBR® Green PCR Kit. No template control (NTC) reactions were also performed (shown by the flat plots). [B] Melting curve analysis, demonstrating high PCR specificity.|QuantiTect Primer Assays enable significant time savings over homemade methods.|
Performance

 QuantiTect Primer Assays在很大模板浓度范围内确保高效定量分析RNA转录本(参见"Reproducible real-time RT-PCR"的A部分)。应用QuantiFast SYBR Green Kit或其他QIAGEN SYBR Green试剂盒可确保精确的定量分析。该试剂盒可避免形成PCR人工产物(参见"Reproducible real-time RT-PCR"的B部分)。

配合QuantiFast、QuantiTect、Rotor-Gene或FastLane SYBR Green Kit(用于两步或一步法RT-PCR的预混液)的分析方法与探针法分析相比,CT值可能更低(参见"Superior sensitivity in real-time RT-PCR")。这得益于试剂盒的高PCR特异性,以及SYBR Green染料与每个PCR产物的多重结合。即使处理低丰度转录本也可进行高灵敏度的定量分析。

Principle
应用QuantiTect Primer Assays无需反复验证real-time RT-PCR引物(参见"Time savings in development of real-time RT-PCR assays")。可用于一步和两步法定量RT-PCR。只需登陆QIAGEN GeneGlobe,选择和订购靶基因的引物对。引物对覆盖所有人类、小鼠、大鼠、果蝇、拟南芥和其他各类生物基因。QuantiTect Primer Assays配合QuantiFast、QuantiTect、Rotor-Gene或FastLane SYBR Green Kit(用于两步或一步法RT-PCR的预混液)能提供可靠的结果。
Procedure

每个QuantiTect Primer Assay包含特定基因的正反引物对,以冻干粉形式提供。在pH8.0的条件下经重悬获得10倍稀释的引物溶液,然后可加入QuantiFast、QuantiTect、Rotor-Gene或FastLane试剂盒提供的预混液,进行SYBR Green法检测。将含有引物的预混液装入PCR反应管或孔板,然后加入单个模板样本。

QuantiTect Primer Assays与表中试剂盒配合使用,可获得可靠的结果。

可与QuantiTect Primer Assays共同使用的两步法和一步法定量RT-PCR试剂盒
两步法qRT-PCR一步法qRT-PCR
QuantiFast SYBR Green PCR Kit QuantiFast SYBR Green RT-PCR Kit
Rotor-Gene SYBR Green PCR Kit Rotor-Gene SYBR Green RT-PCR Kit
QuantiTect SYBR Green PCR Kit QuantiTect SYBR Green RT-PCR Kit

QuantiTect Primer Assays适用于检测RNA。当无法避免基因组DNA污染时,可使用 QuantiTect Reverse Transcription Kit或FastLane Cell cDNA Kit制备cDNA。这两种试剂盒在合成cDNA的同时,可去除基因组DNA污染。

命名 

基因的 QuantiTect Primer Assay均以数字命名区分,数字位于产品名中基因符号之后。更多详情请参照GeneGlobe FAQs。

Applications

QuantiTect Primer Assays适用于利用SYBR Green检测技术、一步或两步法定量RT-PCR进行基因表达分析。非常适合于以下应用:

  • 验证siRNA介导的基因沉默
  • 验证微阵列结果
  • 高通量筛查
Feature
Specifications
Applications Gene expression analysis
Detection SYBR Green I
Format Lyophylized primer set
Kit for this application on the instrument QuantiFast SYBR Green PCR or RT-PCR Kit, QuantiTect SYBR Green PCR or RT-PCR Kit
Real-time or endpoint Real-time
Species Human, mouse, rat, drosophila arabidopsis, chicken, dog, nematode, and zebrafish

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Articles
1
QuantiTect Primer Assays for housekeeping genes can be used as endogenous controls in qRT-PCR.
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Kit Handbooks
1
For genomewide, ready-to-use real-time RT-PCR assays using SYBR Green detection
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