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Pierce™ Magnetic RNA-Protein Pull-Down Kit (Thermo Scientific™)

The Thermo Scientific Pierce Magnetic RNA-Protein Pull-Down Kit provides researchers with a streamlined, robust method to enrich protein-RNA interactions using end-labeled RNA as the bait.

Features of the Magnetic RNA-Protein Pull-Down Kit:

Direct – capture ribonucleoprotein complexes directly with end-labeled RNA; does not use or require antibodies for pull-down
Easy to use – no spin cups or centrifugation necessary for the enrichment of RBP; procedure streamlined for minimal hands-on time (less than 3 hours) after RNA labeling reaction
Flexible – use in vitro transcribed RNA or synthetic RNA for labeling of various lengths and complexity; proteins successfully enriched using endogenous, over-expressed, and in vitro translated lysates
Specific – low bead background; unrelated RNA or mutated RNA does not significantly enrich specified RBPs
Economical – less expensive than purchasing commercially synthesized end-labeled RNA, magnetic beads, and reagents separately
Complete – contains both labeling and enrichment modules with buffers necessary for assay; positive control RNA, negative control RNA, and RBP antibody included

The Magnetic RNA-Protein Pull-Down Kit provides reagents to efficiently enrich RNA Binding Proteins (RBPs) using RNA end-labeled with desthiobiotin and streptavidin magnetic beads. The complete kit contains sufficient reagents for 20 RNA labeling reactions and 20 protein-RNA pull-down assays. This direct enrichment of the protein-RNA interaction provides an alternative to antibody capture of protein-RNA complexes or moieties incorporated into the nucleic acid. An added advantage to the kit is that it includes validated controls for both the labeling and pull-down assay. The kit is amenable to several downstream applications, including Western blotting and Mass Spectrometry (MS).

Includes:
Complete kit contains the Pierce RNA 3'-End Desthiobiotinylation Kit, positive and negative RNA controls, nucleic-acid compatible streptavidin magnetic beads, and buffers for RBP enrichment and elution

Requires:
User-supplied RNA and lysate (experimental sample)

Applications:
• Mutational analysis
• Structure function analysis
• Identification of RNA:Protein interactions
• MS analysis of unknown RNA:protein binding pairs or complexes

Utilizing labeled RNA as bait for RNA-binding protein enrichment is advantageous over antibody enrichment in that it captures the interaction directly. Included in this kit is the Pierce RNA 3'-End Desthiobiotinylation Kit, which uses T4 RNA ligase to attach a single cytidine bisphosphate nucleotide to the 3'-ends of single-stranded RNA. Desthiobiotin may be used for detection or as an elutable affinity handle. The length of the spacer between the nucleotide and desthiobiotin has been optimized for efficient attachment to the beads without compromising the accessibility of the RNA to protein.

The procedure for enrichment of RNA binding proteins has been optimized for ease-of-use. The labeled RNA is first captured to the beads to orient the RNA for protein binding. RNA-bound beads are then equilibrated in Protein RNA Binding buffer before protein lysate is added. After washing, sample can be eluted using either nondenaturing Biotin Elution Buffer or SDS-PAGE loading buffer. Eluted samples are ready for a variety of downstream applications, including Western blotting or Mass spectrometry.

The control system for the pull-down assay utilizes 3'-untranslated region androgen receptor (AR) RNA, poly(A)25 RNA, and mammalian cell lysate. The proximal 3'-untranslated region (UTR) of androgen receptor RNA contains UC-rich regions for HuR and Poly(C) Binding Proteins (CP1 and 2). These RNA binding proteins regulate mRNA stability (HuR) and mRNA turnover and translation (CP1 and 2). The negative control RNA, poly(A)25 RNA, does not contain HuR or poly(C) BP binding sites.

Related Products
Pierce™ Magnetic RNA-Protein Pull-Down Kit

100X Pretreatment Solution (Invitrogen™)

100X Pretreatment Solution is an aqueous buffered solution and an individual component of the ViewRNA™ ISH Tissue Assay Kits, 2-Plex, 24 assays.

TaqMan™ GMO Maize Quantification Kit

The Taqman® GMO Maize Quantification Kit is a food safety testing kit used to determine the number of copies of P35S promoter present in genetically modified organisms compared to the number of copies of maize present in any food or feed sample. The benefits of the Taqman® GMO Maize Quantification Kit over other commercially available products include:

• Efficiency—detection of small DNA fragments through use of Taqman® MGB probes
• Sensitivity—quantification limit of 20 DNA copies; detection limit of 3 DNA copies (maize and P355 systems)
• Flexibility—adapted to processed DNAs

The Taqman® GMO Maize Quantification Kit is effective in detecting the presence of genetically modified organisms through transgenic maize quantification, based on the detection of P35S promoter. The detection success of this promoter is due to this regulatory region being present in most of the transgenic events. In addition, the Taqman® GMO Maize Quantification Kit contains all necessary reagents for quantification and is optimized for use in all (Applied Biosystems®) real-time PCR instruments.

The development and manufacture of this kit occurs through a partnership with IMEGEN (Instituto de Medicina Genomica), which has a long history of GMO research and development.

Other related products include:
GMO Extraction Kit
TaqMan® GMO Screening Kit
Taqman® Roundup Ready Soya Quantification Kit

MAXIscript™ SP6 Kit with Manual (Invitrogen™)

The Ambion® MAXIscript® SP6 In Vitro Transcription Kit synthesizes RNA probes in just 10 minutes for use in ribonuclease protection assays, in situ hybridization, and blot hybridizations. Each kit contains sufficient reagents for 100 reactions.

Features:

• High yields of high specific activity, full-length RNA probes in just 10 minutes
• Ideal probe synthesis kit for nuclease protection assays, northern and Southern blots, and in situ hybridization
• Includes TURBO DNase™
• All kits contain quality-tested DNase/RNase-free reagents

The MAXIscript® In Vitro Transcription Kit synthesizes RNA probes with specific activities reaching 1 × 109 cpm/µg in just 10 minutes. MAXIscript® Kits are very efficient at synthesizing full-length probes, even at limiting nucleotide concentrations, making them the perfect companion to ribonuclease protection assays. MAXIscript® Kits are also an excellent choice for the synthesis of nonisotopically labeled probes. Labeling can be accomplished post-transcriptionally by using Ambion's BrightStar® Psoralen-Biotin Labeling Kit, or during transcription by incorporating modified nucleotides into the reaction. MAXIscript® Kits can also be used for the synthesis of modest amounts of unlabeled RNA (2–6 µg/20 µl reaction) of up to 2 kb.

MEGAscript™ T7 Transcription Kit (Invitrogen™)

The MEGAscript® T7 Kit is an ultra-high yield in vitro transcription kit. The high yields are achieved by modifying typical transcription reaction conditions so that very high nucleotide concentrations can be effectively used. The T7 Enzyme Mix and the 10X Reaction Buffer are specifically calibrated for each lot and RNA polymerase.

Using the MEGAscript® T7 Kit
The MEGAscript® T7 Kit contains in vitro transcription reaction components and a control template. The kit will yield a total of 3–5 mg of RNA (approximately 100 µg of RNA or more per reaction) from the control template supplied with the kit. This corresponds to 400–650 moles of RNA for each mole of template. Smaller templates typically yield a lower mass and a higher molar yield of product. Novel transcription reaction conditions and a patented, high-yield technology allow the synthesis of 10–50 times the amount of RNA produced by conventional transcription reactions.

Applications
The MEGAscript® T7 Kit is intended for the synthesis of large amounts of unlabeled or low specific activity RNA for a variety of uses, including in vitro translation, antisense/microinjection studies, and isolation of RNA binding proteins. In large-scale transcription reactions, the concentration of all 4 nucleotides is high, well above the Km for the enzyme. The MEGAscript® T7 Kit typically yields over 10 times more RNA than conventional in vitro transcription reactions. The kit is not recommended for synthesis of high specific activity probes.

Accessory products
MessageAmp™ aRNA amplification kits are powered by MEGAscript® technology and have been developed especially for the amplification of cRNA prior to array analysis. To purify in vitro transcription products from free nucleotides, buffer components, and enzymes, we recommend using the MEGAclear™ Kit (Cat. No. AM1908) and the MEGAclear™-96 Kit (Cat. No. AM1909).

Wash Buffer Components (1 and 2) (Invitrogen™)

Wash Buffer Components (1 and 2) is an individual component of the ViewRNA™ ISH Tissue Assay Kits, 1-Plex, QVT0050. It is an aqueous buffered solution containing detergent, and can be used for 24 assays (108 mL). Store at -15°C to -30°C.

mirVana™ miRNA Detection Kit with manual (Invitrogen™)

The Ambion® mirVana™ miRNA Detection Kit provides a fast and sensitive method for detecting small RNAs. The assay is 100–500 times more sensitive than Northern analysis, it is able to detect as little as 10 attomoles (10-17 mol) of target RNA. Each kit includes sufficient reagents for 100 reactions.

• Sensitive—detect miRNA or siRNA in as little as 50 ng total RNA
• Specific—extremely low backgrounds
• Simple and fast—single-tube procedure eliminates the need to transfer to membrane for hybridization
• Multiple target detection—detect multiple small RNAs and mRNAs in the same sample

Rapid Procedure Based on Solution Hybridization
The mirVana™ miRNA Detection Kit is based on a simple solution hybridization principle where the RNA sample is simply mixed with radiolabeled RNA probe(s) and hybridized. Unhybridized RNA and excess probe are removed by a digestion step and the hybridized, protected RNA fragments are then recovered using a patented single-step technology for simultaneous ribonuclease inactivation and nucleic acid precipitation. RNA samples are then resuspended and analyzed on a denaturing polyacrylamide gel.

Simultaneous Detection of siRNA Expression and Target Gene Knockdown
Because multiple RNA species of the same or differing size can be detected in the same sample, the mirVana™ miRNA Detection Kit is ideal for correlating siRNA expression levels with target mRNA knockdown. An example of this application is shown in the accompanying data, which demonstrates that GAPDH mRNA levels were reduced in cells expressing a GAPDH siRNA, but not in cells expressing a control siRNA.

Accessory Products:
The mirVana™ miRNA Probe Construction Kit (SKU#AM1550) is available for the preparation and labeling of probes by in vitro transcription and the mirVana™ Probe & Marker Kit is available for 5' end labeling of oligonucleotide probes and the included Decade Markers (SKU#AM1554).

ViewRNA™ ISH Cell Assay Kit (Invitrogen™)

The ViewRNA™ ISH Cell Assay is a direct fluorescence RNA in situ hybridization method that enables the simultaneous detection of one to four RNA targets (either mRNA or non-coding RNA) at single copy sensitivity and single cell resolution using fluorescence microscopes or high-content imagers. Unlike traditional FISH techniques which are generally limited by high background and low sensitivity, the assay uses proprietary chemistry for the target specific probe sets (RNA FISH probes) and branch DNA signal amplification (bDNA) for detection of specific signal. The fluorescence RNA in situ hybridization assay has four main steps: sample preparation, target hybridization, signal amplification, and detection. This ViewRNA ISH assay kit can detect up to three RNA but can be combined with the ViewRNA™ 740 Module to add the fourth probe.

Required additional components:
10X PBS (cat. no. QVC0508)
ViewRNA™ - Detergent Solution QC (cat. No. QVC0509)
ViewRNA™ - Wash Buffer Set (cat. no. QG0507)
ViewRNA™ ISH Cell Accessory Kit (cat. no. QVC0700) intended to provide many of the required components, not supplied in the reagent kit, in order to perform the assay.
ViewRNA™ Probe Sets are not included in the assay and are designed for use with the ViewRNA ISH Cell Assays. Visit our website to view a complete listing of over 6,500 synthesized Probe Sets. By request, new Probe Sets can be designed and synthesized in less than two weeks with no additional costs.

To add the 4th RNA probe:
ViewRNA™ ISH Cell 740 Module (cat. no. QVC0200) designed to be used in conjunction with ViewRNA™ ISH Cell Assay and allows analysis of an additional RNA target in the 740 channel (AlexaFluor 750) See the package insert for a complete list of materials provided in the kit.

Reported Applications
Microscopy

MAXIscript™ T7 Transcription Kit (Invitrogen™)

The Ambion® MAXIscript® T7 In Vitro Transcription Kit synthesizes RNA probes in just 10 minutes for use in ribonuclease protection assays, in situ hybridization, and blot hybridizations. Each kit contains sufficient reagents for 30 reactions.

Features:

• High yields of high specific activity, full-length RNA probes in just 10 minutes
• Ideal probe synthesis kit for nuclease protection assays, northern and Southern blots, and in situ hybridization
• Includes TURBO DNase™
• All kits contain quality-tested DNase/RNase-free reagents

The MAXIscript® In Vitro Transcription Kit synthesizes RNA probes with specific activities reaching 1 × 109 cpm/µg in just 10 minutes. MAXIscript® Kits are very efficient at synthesizing full-length probes, even at limiting nucleotide concentrations, making them the perfect companion to ribonuclease protection assays. MAXIscript® Kits are also an excellent choice for the synthesis of nonisotopically labeled probes. Labeling can be accomplished post-transcriptionally by using Ambion's BrightStar® Psoralen-Biotin Labeling Kit, or during transcription by incorporating modified nucleotides into the reaction. MAXIscript® Kits can also be used for the synthesis of modest amounts of unlabeled RNA (2–6 µg/20 µl reaction) of up to 2 kb.

ViewRNA™ Tissue Assay Core Kit (Invitrogen™)

The ViewRNA Tissue Assay Core Kit, when combined with a ViewRNA Tissue probe set (purchased separately), enables the detection of a single mRNA target in tissue sections using the Fast Red substrate. The ViewRNA Tissue Assay Blue Module enables the detection of a second mRNA target using the Fast Blue substrate and must be used in combination with the Core Kit and two probes sets in order to run a 2-plex assay. Each kit contains sufficient reagents to process a minimum of six slides at a time. ViewRNA TYPE 1 probe sets are for use with the Fast Red substrate (Core Kit), while ViewRNA TYPE 6 probe sets are for use with the Fast Blue substrate (Blue Module).

Learn more about and order ViewRNA Tissue probe sets ›

ViewRNA Tissue assays are RNA in situ hybridization (ISH) assays for the reliable detection of one or two mRNA targets within tissue sections. Visualize any gene in any tissue with single-copy sensitivity and no radioactivity. And in case your needs are not met by one of our 6500 catalog probe sets, we have decades of bioinformatics expertise and will design custom probe sets to any mRNA target of interest, free of charge.

With ViewRNA Tissue assays, you can achieve:
• Robust, single mRNA molecule detection in tissue sections
• Accuracy and specificity inherent to branched DNA amplification technologies
• Simultaneous detection of two RNA targets

The ViewRNA Tissue Assay is compatible with formalin-fixed, paraffin-embedded (FFPE) tissue or OCT (optimal cutting temperature) embedded frozen tissue sections. Whereas traditional ISH technologies often amplify both signal and background, ViewRNA assays use branched DNA (bDNA) technology to amplify target-specific signal and achieve superior signal-to-noise ratios.

Fast Blue Substrate (20 mL Blue Buffer + 420 uL each Blue Reagents 1-3) (Invitrogen™)

Fast Blue Substrate (20 mL Blue Buffer + 420 uL each Blue Reagents 1-3) is an individual component of the ViewRNA™ ISH Tissue Assay Kits, 2-Plex, 24 assays. It is a blue precipitating substrate with components 1 -3 used for the detection of alkaline phosphatase activity.

RapidFinder™ Quant Multi-Meat Set

The RapidFinder™ Quant Multi-Meat Set, when used with one or more of the RapidFinder™ Meat ID kits listed below, enables the determination of the percentage of a target meat species with respect to total meat in food and feed samples. Using the included standards, the kit enables quantification of as little as 0.05% target species DNA as compared to other meat DNA in only a few hours.

The RapidFinder Quant Multi-Meat Set is designed to be used in combination with one or more of the following meat species-specific ID kits (available separately):

NameCatalog Number
RapidFinder™ Beef ID KitA24391
RapidFinder™ Pork ID KitA24392
RapidFinder™ Equine ID KitA15570
RapidFinder™ Poultry ID KitA24397
RapidFinder™ Chicken ID KitA24393
RapidFinder™ Turkey ID KitA24394


• Simple-real-time PCR based assay, no electrophoresis needed
• Flexible-compatible with any food or feed sample, both raw and processed
• Reliable-Includes multi-meat DNA quantification standard
• Fast-from sample to result in the same day

How It Works
The method relies on quantitative real-time PCR. First, a standard curve is created by serially diluting the included multi-meat DNA standard. Then assays for the target species DNA (from one of the kits listed above) and for all meat DNA (from the RapidFinder™ Quant Multi-Meat Set) are run side-by-side on both the food sample DNA and the DNA standards. This approach enables rapid and accurate quantification and reporting of the percentage of target meat species as compared to total meat in a sample. Included in each reaction mix is an internal positive control detected in the VIC® dye channel, which is used to verify the absence of PCR inhibitors.

The RapidFinder™ Quant Multi-Meat Set is compatible with Applied Biosystems® StepOne™, StepOnePlus™, 7500, 7500 Fast, and 7300 real-time PCR systems.

Sample Preparation
DNA sample preparation for the RapidFinder™ meat species identification and quantification workflow is accomplished with the easy-to-use GMO Extraction Kit (Cat. No. 4466336). Using 10 g of sample rather than the smaller sample sizes accommodated by most other DNA isolation procedures minimizes sampling errors. The GMO Extraction Kit is compatible with raw and processed meat, as well as composite food samples.

Click-iT™ RNA Alexa Fluor™ 594 Imaging Kit (Invitrogen™)

The Click-iT® RNA Alexa Fluor® 594 Imaging Kit enables detection of global RNA transcription temporally and spatially in cells and tissues (PNAS (2008) 105:15779-84).The ability to detect newly synthesized RNA or changes in RNA levels resulting from disease, environmental damage or drug treatments is an important aspect of toxicological profiling. Utilizing an alkyne-modified nucleoside, 5-ethynyl uridine (EU), and powerful click chemistry, newly synthesized RNA can be detected without the use of radioactivity or antibodies with a simple, two-step procedure. In step one, the alkyne-containing nucleoside is fed to cells or animals and actively incorporated into nascent RNA. The small size of the tag enables efficient incorporation of the modified nucleoside into RNA, but not into DNA. Detection utilizes the chemoselective ligation or “click" reaction between and azide and an alkyne where the modified RNA is detected with a corresponding azide-containing dye. With its dimunitive “footprint", the Click-iT® detection molecule can easily penetrate complex samples and leaves open the possibility of multiplex analyses with other probes, including antibodies for the detection of RNA-interactive proteins for deeper biological insights.

FISH Tag™ RNA Green Kit, with Alexa Fluor™ 488 dye (Invitrogen™)

FISH Tag™ RNA Kits provide a complete workflow solution for fluorescence in situ hybridization (FISH) applications. Each kit provides all of the reagents needed for probe synthesis, labeling, and purification, as well as for imaging the labeled specimen. Choose from FISH Tag™ RNA Kits in two Alexa Fluor® colors or get the FISH Tag™ RNA Multicolor Kit, which contains four spectrally distinct dyes to allow simultaneous localization of multiple sequence-specific RNA species in cells, tissues, and whole embryo mounts.

FISH Tag™ Kit Specifications:
• Dye (Ex/Em): Alexa Fluor® 488 (492/520 nm)
• Comprehensive kit includes 16 reagents
• Comes with detailed protocol to help ensure your success


Complete Workflow Solution for FISH
FISH Tag™ Kits employ a two-step labeling technology—nick translation to enzymatically incorporate an amine-modified nucleotide (aminoallyl dUTP) followed by chemical labeling with Alexa Fluor® dyes. PureLink® nucleic acid purification technology is used for fast and efficient purification of the labeled probe, and SlowFade® Gold antifade reagent is added to preserve the fluorescent signal during imaging.

More Options for Nucleic Acid Labeling
To review various options for nucleic acid labeling (including DNA and RNA FISH), review Labeling Oligonucleotides and Nucleic Acids—Section 8.2 in the Molecular Probes® Handbook.

For Research Use Only. Not intended for human or animal therapeutic or diagnostic use.

RapidFinder™ Beef ID Kit

The RapidFinder™ Beef ID Kit enables the simple and reliable detection of beef in food and feed samples. The method works by detection of bovine, or Bos taurus, DNA by real-time PCR. Unlike ELISA-based methods, when used with the GMO Extraction Kit (Cat. No. 4466336) for sample preparation, the RapidFinder™ Beef ID Kit detects both raw and processed beef and provides sensitivity down to 0.01% bovine DNA. The included positive control, which comprises 0.1% bovine DNA, can be used as a reference to set the threshold of detection of the assay.

• Simple-real-time PCR based assay, no electrophoresis needed
• Flexible-bovine DNA detected in any food or feed sample, both raw and processed
• Reliable-Internal Positive Control (IPC) provides process-control check for PCR inhibitors
• Fast-from sample to result in the same day

How the RapidFinder™ Beef ID Kit Works
Bovine DNA detection is accomplished by real-time PCR using the supplied primer/probe set specific for mitochondrial DNA sequence specific to Bos taurus. The target sequence, detected in the FAM™ dye channel, is not only specific for beef, but also stable. Also included in each reaction mix is an internal positive control detected in the VIC® dye channel, which is used to verify the absence of PCR inhibitors.

By performing a separate reaction with the included positive control, users are able to verify the system and assay are working correctly. More importantly, it enables easy data interpretation. The positive control is provided as 0.1% bovine DNA in a background of fish DNA, and by comparing sample results to the control results, a threshold of detection is provided.

The assay is compatible with Applied Biosystems® StepOne™, StepOnePlus™, 7500, 7500 Fast, and 7300 real-time PCR systems.

Sample Preparation
DNA sample preparation for the RapidFinder™ Beef ID Kit workflow is accomplished with the easy-to-use GMO Extraction Kit (Cat. No. 4466336). Using 10 g of sample rather than the smaller sample sizes accommodated by most other DNA isolation procedures minimizes sampling errors. The GMO Extraction Kit is compatible with raw and processed meat, as well as composite food samples.

Quantification
If a positive result is obtained with the RapidFinder™ Beef ID Kit, most labs will go on to quantify the amount of bovine DNA in the sample. This can be accomplished using the RapidFinder™ Beef ID Kit in conjunction with the RapidFinder™ Quant Multi-Meat Set (Cat. No. A24399).

Part of a Suite of Solutions For Meat Species Detection
The RapidFinder™ Beef ID Kit is just one of our many kits available for the sensitive detection of various meat species in food and feed samples. Other RapidFinder™ meat species identification kits available include those for the detection of pork, horse, sheep, ruminant, chicken, turkey, and poultry.

The RapidFinder™ Meat Species Identification kits are made available through a partnership with IMEGEN (Instituto de Medicina Genomica).