This procedure uses denatured RNase T1 to digest RNA. RNase T1 cleaves single-stranded RNA residues of guanosine 3 '. Partial digestion of 3 'or 5 like' RNA with this enzyme thus produces a scale of residue G.
The 1X rna synthesis Buffer used in the procedure containing 7 M urea denaturing contribute to RNA secondary structure. To perform the procedure diluted the RNA ends at least five times in buffer sequencing. The sample is denatured at 50 ° C, RNA and heatedthen incubated with RNase T1 at room temperature.
rna synthesis
In the process, two different concentrations of RNase T1 can be used. Tube # 1 contains RNase and represents a negative control. Non-Bands in length in the tube are fission products within the RNA sample. These bands will also be present in samples of nuclease treatment, should be ignored in its analysis. T1 Tubes # 2 and # 3 with two different concentrations of RNase. Normally, at least one of the reactionscreate a ladder with digestion by each of the guanosines in the rna synthesis . Further enzyme dilution may be necessary to achieve the optimum scale digestion.
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