Analyze RNA with Reagents You Can Trust
RNA size markers come in three different size ranges (0.5-9 kb, 100-1000 bases, and 10-150 bases) and can be detected through autoradiography, chemiluminescence, or fluorescence. You can also rely on Ambion to provide molecular biology grade reagents for making and running electrophoretic gels (both agarose and acrylamide). As described below, several of our solutions have been optimized for increased laboratory safety, ease-of-use, or nucleic acid fractionation.
Markers for mRNA Analysis
* RNA Millennium Markers (500-9000 nt)
Millennium Markers are a mixture of ten discrete RNA transcripts of sizes (0.5, 1, 1.5, 2, 2.5, 3, 4, 5, 6, and 9 kilobases, each with a 20 base poly(A) tail) that can be easily detected on a 1% denaturing agarose gel. The markers may be stained with ethidium bromide during or after electrophoresis, or they can be end-labeled using T4 polynucleotide kinase (e.g. using Ambion's KinaseMax™ Kit, Cat #1520). RNA Millennium Markers are provided in aqueous solution or deionized formamide.
* BrightStar® Biotinylated RNA Millennium Markers (500-9000 nt)
BrightStar Biotinylated Millennium Markers are a mixture of unlabeled and biotinylated Millennium Markers that can be stained using ethidium bromide and detected nonisotopically using any biotin detection system (e.g. BrightStar® BioDetect™ Nonisotopic Detection Kit, Cat #1930).
* Millennium Marker Probe Template
The Millennium Marker Probe Template offers an alternative to end-labeling the Millennium Markers directly. When transcribed in vitro with SP6, T7, or T3 RNA phage polymerases, the Marker Probe Template (a cloned 0.43 kb fragment) produces an RNA complementary to the Millennium Markers and will hybridize with all ten Millennium Marker transcripts on Northern blots.
Markers for Small RNA Analysis
RNA Century Size Markers (100-500 nt)
RNA Century Markers include five transcripts of 100, 200, 300, 400, and 500 bases. Century Markers come premixed to give equivalent band staining on 5% polyacrylamide/8 M urea gels. These markers can also be radioactively end-labeled.
RNA Century-Plus Size Markers (100-1000 nt)
RNA Century-Plus Markers have two additional, longer RNA transcripts compared to the RNA Century Size Markers (100, 200, 300, 400, 500, 750, and 1000 bases). Like the Century Markers, the Century-Plus Markers come premixed to give equivalent band staining on 5% polyacrylamide/8 M urea gels and can be radioactively end-labeled.
Century Marker Template and Century-Plus Marker Template
These two sets of templates allow researchers to synthesize Century Marker or Century-Plus Marker RNAs that are labeled with radioisotopes, biotin, or other modified nucleotides (e.g. using Ambion's MAXIscript® Kit, Cat #1308-1326). Each set of templates contains a mixture of linearized plasmids that is ready for in vitro transcription reactions using T7 RNA polymerase. Under standard reaction conditions, the bands created by each size of transcript will have essentially equal ethidium bromide staining intensity.
BrightStar Biotinylated RNA Century and Century-Plus Markers (100-500 nt or 100-1000 nt)
BrightStar Biotinylated RNA Century and Century-Plus Markers can be visualized with ethidium bromide staining and/or by a biotin detection system (e.g. Ambion's BrightStar BioDetect Nonisotopic Detection Kit, Cat #1930). Both BrightStar Biotinylated RNA Century and Century-Plus Markers are premixed so each band gives equivalent signal intensity and are useful as size standards in nonisotopic nuclease protection assays.
Decade Markers (10-150 nt)
The Decade Marker System allows researchers to produce a set of radiolabeled RNA molecular weight markers of 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, and 150 bases. This system contains all the reagents, except for [gamma-32P]ATP, needed to end-label and cleave a gel-purified transcript to generate the RNA ladder. The marker is ready for use in less than one hour, and a single reaction can be used for 5-20 experiments.
Marker for the Agilent® 2100 Bioanalyzer
The RNA 6000 Ladder is a set of six RNA transcripts with lengths of 0.2, 0.5, 1.0, 2.0, 4.0, and 6.0 kb. This ladder is designed for use with the Agilent 2100 bioanalyzer and RNA 6000 LabChip® Kits. The Agilent 2100 bioanalyzer is a microfluidics-based platform for analysis of nucleic acids, proteins, and cells that requires minimal sample for analysis (e.g. 5 ng total RNA) and is accurate, rapid, and safe.
Reagents for Agarose Gel Electrophoresis
Ambion has two types of agarose that can both be used in formaldehyde or glyoxal denaturing gels.
* Agarose-LE™ General Use
Agarose-LE is suitable for routine electrophoresis of nucleic acid or protein. It has a low electroendosmosis value (EEO-mr 0.103) ensuring high electrophoretic mobility of nucleic acids.
* Agarose-HR™ High Resolution
Agarose-HR is optimized to separate nucleic acids that are 50 to 1000 bp in length. It is useful for sizing PCR fragments, small DNA fragments generated by restriction enzyme digestion, and double-stranded RNA fragments. Nucleic acid resolution is enhanced because of low background fluorescence with ethidium bromide.
Gel Loading Solutions
* NorthernMax® Formaldehyde Load Dye
This ready-to-use solution is added to RNA samples (3 parts solution : 1 part sample) and heated briefly before electrophoresis. This solution already contains xylene cyanol and bromophenol blue; ethidium bromide can be added to the samples, if desired.
* NorthernMax®-Gly Sample Loading Dye
NorthernMax-Gly loading dye is an improved formulation over traditional recipes, and can be used for RNA sample denaturation in any glyoxal gel protocol. The volume ratio of solution to sample is lower than in published protocols, so sample precipitation prior to gel loading is usually not required. Bromophenol blue and ethidium bromide are premixed into the solution.
* NorthernMax® 10X Running Buffer
NorthernMax Running Buffer is a 10X MOPS-based solution that contains sodium acetate and EDTA. MOPS is a zwitter-ionic buffer that helps to reduce heat generation while allowing timely nucleic acid separation on denaturing formaldehyde agarose gels.
* NorthernMax® 10X Denaturing Gel Buffer
This is an improved formulation for use in denaturing formaldehyde/MOPS agarose gels. Because this buffer makes the gels less brittle than conventional gels, these gels are much easier to handle.
* NorthernMax®-Gly 10X Gel Prep/Running Buffer
When used with NorthernMax-Gly Sample Loading Dye, the NorthernMax-Gly Running Buffer can improve resolution of RNA compared to standard glyoxal or formaldehyde systems.
Reagents for Acrylamide Gel Electrophoresis
Ambion's liquid Acrylamide/Bis solutions are ready for use, so the dust, inhalation, and contact hazards associated with weighing and preparing acrylamide and bis-acrylamide is reduced. Concentrations are carefully controlled, and all solutions are micro-filtered prior to packaging.
* 6% PAGE/6 M Urea Solution (19:1 Acrylamide/Bis with Urea)
When used as directed, the final gel will consist of 6 M urea, 6% acrylamide (19:1 acrylamide/bis-acrylamide) in 1X TBE at pH 8.3, which is ideal for use in all denaturing polyacrylamide gel electrophoresis of nucleic acids (e.g. ribonuclease protection assays, gel purification of RNA probes, PCR fragment analysis, sequencing).
* Acrylamide/Bis 19:1 40% (w/v) solution
For increased flexibility, Ambion offers the convenience of a 19:1 acrylamide/bis-acrylamide solution that can be used for nondenaturing gels or can be customized (e.g. urea or acrylamide concentration) for your specific experiment.
Gel Loading Solutions
* NorthernMax® Formaldehyde Load Dye
In addition to agarose gels, this loading buffer can actually be used with polyacryl-amide gels and does not require urea. It may be necessary to stain the gel after electrophoresis, even if ethidium bromide was added to the buffer.
* Gel Loading Buffer II--Denaturing PAGE
This buffer is a 1-2X solution of 95% formamide, 18 mM EDTA, and 0.025% SDS, and contains xylene cyanol and bromophenol blue, making it an ideal gel loading buffer for polyacrylamide/urea gel electrophoresis. Compared to many sample loading buffers, it is also less viscous, so less sample will cling to tips and tubes.
10X TBE Running Buffer
10X TBE running buffer is supplied as packages (1 L of 10X TBE per package) of ultra pure, molecular biology grade powder. The buffer will contain 89 mM Tris-Borate and 2 mM EDTA when diluted to the 1X working solution.