New England Biolabs, N0364S, Low Range ssRNA Ladder

Related Categories RNA Markers & Ladders,, RNA Markers and Ladders Specification Storage Buffer 20 mM sodium citrate 1 mM EDTA pH 6 @ 25°C FAQ Q: What types of gels are recommended for ssRNA and dsRNA ladders? (N0363, N0362, N0364) A: In general, when working with single-stranded RNA molecules, one should use a denaturing gel, a denaturing sample loading buffer/dye containing either formamide or urea, and SYBR Gold for staining. The SYBR Gold stain binds preferentially to single-stranded molecules and therefore it give good visualization of the bands after gel electrophoresis. In some instances, when ssRNA molecules are large in size such as the ssRNA Ladder (N0362S), one can use a native gel and stain the gel with ethidium bromide. But keep in mind that the ssRNA samples must always be prepared using a denaturing sample loading buffer/dye containing either formamide or urea. When dealing with double-stranded RNA molecules, one only needs to use a native gel, a non-denaturing sample loading buffer/dye, and stain the gel with either ethidium bromide, SYBR Green or SYBR Safe. Q: What types of gel and staining dyes can I use for NEB’s Low Range ssRNA Ladder (N0364S)? A: The prepared Low Range ssRNA Ladder (N0364S) can be run either on a native 2.0% agarose-TBE or a denaturing 6% TBE-Urea gel (Life Technologies). The gel can be stained using either SYBR Gold or ethidium bromide. Q: Why the bands of the Low Range ssRNA Ladder on my 2% agarose-TBE gel are not resolved well? A: You must be sure to do the denaturation of the ladder properly. After adding 2µL of Low Range ssRNA Ladder to 38µL of RNA Loading Dye, you must incubate this mixture at 90˚C for 3-5 minutes or at 70˚C for 10 minutes and chill it on ice for 2 minutes before loading. Q: Why do I have extra bands of the Low Range ssRNA Ladder on my denaturing 6% TBE-Urea gel? A: Having extra bands on a denaturing gel is an indication that the ladder was not denatured properly and/or you overloaded the ladder. Q: Will the Low Range ssRNA Ladder run on 10% TBE-Urea and 15% TBE-Urea gels? A: Yes. When using denaturing gels of high % polyacrylamide to run the Low Range ssRNA Ladder, you should use lesser amount of ladder than what is recommended in the protocol for running a 6% TBE-Urea. For example, you only need 0.03µL-0.06µL of ladder per lane on a Novex 10% TBE-Urea (10-well) gel or ≤0.03µL of ladder per lane on a Novex 15% TBE-Urea (12-well) gel. Q: What is the concentration of RNA in the ssRNA Ladder (NEB #N0362) and Low Range ssRNA Ladder (NEB #N0364)? A: Since the RNA ladders are designed to be size standards and not meant for quantitation, we do not publish the concentration of these products. The release criteria for the ladders is visual inspection of the electrophoretic pattern. Even though we try our best to keep the ladder profile and band intensity consistent, the RNA concentration may vary between batches. Due to this we do not recommend these products for quantitation purposes.