qScript One-Step SYBR Green RT-qPCR

1-step SYBR simplicity

Features & Benefits

  • Sensitive RNA detection with performance-engineered, RNase H (+) M-MLV reverse transcriptase
  • Stringent ultra-pure antibody hot start ensures precise target amplification
  • Flexible buffer chemistry supports either conventional or accelerated thermal cycling conditions

 

qScript One-Step SYBR Green RT-qPCR Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention or treatment of a disease.

Description

The qScript One-Step SYBR Green RT-qPCR Kit is a convenient and highly sensitive solution for quantitative RT-PCR of RNA templates (RT-qPCR) using SYBR Green I dye detection and gene-specific primers. cDNA synthesis and PCR amplification are carried out in the same tube without opening between procedures.The proprietary reaction buffer has been specifically formulated to maximize activities of both reverse transcriptase and Taq DNA polymerase while minimizing the potential for primer-dimer and other non-specific PCR artifacts. This reagent is compatible with both fast and standard qPCR cycling protocols. Precise amplification is essential for successful RT-qPCR with SYBR Green I technology since this dye binds to all dsDNA generated during amplification. This 1-step reagent contains ultra-pure AccuStart™ hot start Taq DNA polymerase that is completely arrested prior to the initial PCR denaturation step. Upon heat activation at 95°C, the antibodies are rapidly and irreversibly denatured, releasing a fully active high-yielding Taq DNA polymerase mutant.

Details

  • Contents
    50X concentrated qScript One-Step Reverse Transcriptase - Optimized 50X formulation of recombinant MMLV reverse transcriptase for one-step RT-PCR. One-Step SYBR Green Master Mix (2X) - 2X reaction buffer containing dNTPs, magnesium chloride, AccuStart Taq DNA polymerase, stabilizers, and SYBR Green I dye Nuclease-free water
  • Storage & Handling
    Store components in a constant temperature freezer at -25°C to -15°C upon receipt. For lot specific expiry date, refer to package label, Certificate of Analysis or Product Specification Form.
  • Instrument Capability
    ROX
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    Low ROX
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    No ROX
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  • Related Resources
    Product Manuals
    Safety Data Sheets (SDS)
    CofA (PSF)
    Publications
    Heterogeneous Nuclear Ribonucleoprotein L is required for the survival and functional integrity of murine hematopoietic stem cells : Scientific Reports
    Marie-Claude Gaudreau, Scientific Reports - 2016
    Abstract
    The proliferation and survival of hematopoietic stem cells (HSCs) has to be strictly coordinated to ensure the timely production of all blood cells. Here we report that the splice factor and RNA binding protein hnRNP L (heterogeneous nuclear ribonucleoprotein L) is required for hematopoiesis, since its genetic ablation in mice reduces almost all blood cell lineages and causes premature death of the animals. In agreement with this, we observed that hnRNP L deficient HSCs lack both the ability to self-renew and foster hematopoietic differentiation in transplanted hosts. They also display mitochondrial dysfunction, elevated levels of γH2AX, are Annexin V positive and incorporate propidium iodide indicating that they undergo cell death. Lin-c-Kit+ fetal liver cells from hnRNP L deficient mice show high p53 protein levels and up-regulation of p53 target genes. In addition, cells lacking hnRNP L up-regulated the expression of the death receptors TrailR2 and CD95/Fas and show Caspase-3, Caspase-8 and Parp cleavage. Treatment with the pan-caspase inhibitor Z-VAD-fmk, but not the deletion of p53, restored cell survival in hnRNP L deficient cells. Our data suggest that hnRNP L is critical for the survival and functional integrity of HSCs by restricting the activation of caspase-dependent death receptor pathways.
    The liver-specific microRNA-122*, the complementary strand of microRNA-122, acts as a tumor suppressor by modulating the p53/mouse double minute 2 homolog circuitry
    Alina Simerzin, Hepatology - 2016
    Abstract
    The tumor suppressor p53 is a central regulator of signaling pathways that controls the cell cycle and maintains the integrity of the human genome. p53 level is regulated by mouse double minute 2 homolog (Mdm2), which marks p53 for proteasomal degradation. The p53-Mdm2 circuitry is subjected to complex regulation by a variety of mechanisms, including microRNAs (miRNAs). We found a novel effector of this regulatory circuit, namely, miR-122*, the passenger strand of the abundantly expressed liver-specific miR-122. Here, we demonstrate that miR-122* levels are reduced in human hepatocellular carcinoma (HCC). We found that miR-122* targets Mdm2, thus participating as an important player in the p53-Mdm2 circuitry. Moreover, we observed significant negative correlation between levels of miR-122* and Mdm2 in a large set of human HCC samples. In vivo tumorigenicity assays demonstrate that miR-122* is capable of inhibiting tumor growth, emphasizing the tumor-suppressor characteristics of this miRNA. Furthermore, we show that blocking miR-122 in murine livers with an antagomiR-122 (miRNA inhibitor) results in miR-122* accumulation, leading to Mdm2 repression followed by elevated p53 protein levels. Conclusion: We show that miR-122*, the passenger strand of miR-122, regulates the activity of p53 by targeting Mdm2. Importantly, similarly to miR-122, miR-122* is significantly down-regulated in human HCC. We therefore propose that miR-122* is an important contributor to the tumor suppression activity previously attributed solely to miR-122. (Hepatology 2016)
    The impact of the long-distance transport of a BEL1-Like Messenger RNA on Development
    Tian Lin, Original Research - 2016
    Abstract
    BEL1- and KNOTTED1-type proteins are transcription factors from the three-amino-loop-extension superclass that interact in atandem complex to regulate the expression of target genes. In potato (Solanum tuberosum), StBEL5 and its Knox protein partner regulate tuberization by targeting genes that control growth. RNA movement assays demonstrated that StBEL5 transcripts move through the phloem to stolon tips, the site of tuber induction.StBEL5 messenger RNA originates in the leaf, and its movement to stolons is induced by a short-day photoperiod. Here, we report the movement of StBEL5 RNA to roots correlated with increased growth, changes in morphology, and accumulation of GA2-oxidase1,YUCCA1a, and ISOPENTENYL TRANSFERASE transcripts. Transcription of tBEL5 n leaves is induced by light but insensitive to photoperiod, whereas in stolon tips growing in the dark, romoter activity is enhanced by short days. The heterodimer of StBEL5 and POTH1, a KNOTTED1-type transcription factor,binds to a tandem TTGAC-TTGAC motif that is essential for regulating transcription. The discovery of an inverted tandem motifin the tBEL5 romoter with TTGAC motifs on opposite strands may explain the induction of tBEL5 romoter activity in stolon ips under short days. Using transgenic potato lines, deletion of one of the TTGAC motifs from the tBEL5 romoter results in he reduction of GUS activity in new tubers and roots. Gel-shift assays demonstrate BEL5/POTH1 binding specificity to the otifs present in the tBEL5 promoter and a double tandem motif present in the StGA2-oxidase1 promoter. These results suggest that, in addition to tuberization, the movement of StBEL5 messenger RNA regulates other aspects of vegetative development.
    Regulation of fibronectin gene expression in cardiac fibroblasts by scleraxis
    Rushita A. Bagchi, Cell and Tissue Research - 2016
    Abstract
    The glycoprotein fibronectin is a key component of the extracellular matrix. By interacting with numerous matrix and cell surface proteins, fibronectin plays important roles in cell adhesion, migration and intracellular signaling. Up-regulation of fibronectin occurs in tissue fibrosis, and previous studies have identified the pro-fibrotic factor TGFβ as an inducer of fibronectin expression, although the mechanism responsible remains unknown. We have previously shown that a key downstream effector of TGFβ signaling in cardiac fibroblasts is the transcription factor scleraxis, which in turn regulates the expression of a wide variety of extracellular matrix genes. We noted that fibronectin expression tracked closely with scleraxis expression, but it was unclear whether scleraxis directly regulated the fibronectin gene. Here, we report that scleraxis acts via two E-box binding sites in the proximal human fibronectin promoter to govern fibronectin expression, with the second E-box being both sufficient and necessary for scleraxis-mediated fibronectin expression to occur. A combination of electrophoretic mobility shift and chromatin immunoprecipitation assays indicated that scleraxis interacted to a greater degree with the second E-box. Over-expression or knockdown of scleraxis resulted in increased or decreased fibronectin expression, respectively, and scleraxis null mice presented with dramatically decreased immunolabeling for fibronectin in cardiac tissue sections compared to wild-type controls. Furthermore, scleraxis was required for TGFβ-induced fibronectin expression: TGFβ lost its ability to induce fibronectin expression following scleraxis knockdown. Together, these results demonstrate a novel and required role for scleraxis in the regulation of cardiac fibroblast fibronectin gene expression basally or in response to TGFβ.
    Stationary-Phase Mutagenesis in Stressed Bacillus subtilis Cells Operates by Mfd-Dependent Mutagenic Pathways
    Martha Gómez-Marroquín, MDPI - 2016
    Abstract
    In replication-limited cells of Bacillus subtilis, Mfd is mutagenic at highly transcribed regions, even in the absence of bulky DNA lesions. However, the mechanism leading to increased mutagenesis through Mfd remains currently unknown. Here, we report that Mfd may promote mutagenesis in nutritionally stressed B. subtilis cells by coordinating error-prone repair events mediated by UvrA, MutY and PolI. Using a point-mutated gene conferring leucine auxotrophy as a genetic marker, it was found that the absence of UvrA reduced the Leu+ revertants and that a second mutation in mfd reduced mutagenesis further. Moreover, the mfd and polA mutants presented low but similar reversion frequencies compared to the parental strain. These results suggest that Mfd promotes mutagenic events that required the participation of NER pathway and PolI. Remarkably, this Mfd-dependent mutagenic pathway was found to be epistatic onto MutY; however, whereas the MutY-dependent Leu+ reversions required Mfd, a direct interaction between these proteins was not apparent. In summary, our results support the concept that Mfd promotes mutagenesis in starved B. subtilis cells by coordinating both known and previously unknown Mfd-associated repair pathways. These mutagenic processes bias the production of genetic diversity towards highly transcribed regions in the genome. Keywords:
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