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- Q:
Column blockage
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A-1 Cell lysis or homogenization not sufficient
---- Reduce sample usage, increase the amount of lysis buffer, increase homogenization and lysis time.
A-2 Sample amount is too large
---- Reduce the amount of sample used or increase the amount of lysis buffer.
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- Q:
Low RNA yield
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A-1 Insufficient cell lysis or homogenization
---- Reduce sample usage, increase the amount of lysis buffer, increase homogenization and lysis time.
A-2 Sample amount is too large
----Please refer to the maximum processing capacity.
A-3 RNA is not eluted completely from the column
---- After adding RNase-Free water, leave it for a few minutes before centrifuging.
A-4 Ethanol in the eluent
---- After rinsing, centrifuge again and remove the washing buffer as much as possible.
A-5 Cell culture medium is not completely removed
---- When collecting cells, please make sure to remove the culture medium as much as possible.
A-6 The cells stored in RNAstore are not effectively centrifuged
----RNAstore density is greater than the average cell culture medium; so the centrifugal force should be increased. It is suggested to centrifuge at 3000x g.
A-7 Low RNA content and abundance in the sample
---- Use a positive sample to determine if the low-yield is caused by the sample.
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- Q:
RNA degradation
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A-1 The material is not fresh
---- Fresh tissues should be stored in liquid nitrogen immediately or immediately put into the RNAstore reagent to ensure the extraction effect.
A-2 Sample amount is too large
---- Reduce sample amount.
A-3 RNase contamination
----Although the buffer provided in the kit does not contain RNase, it is easy to contaminate RNase during extraction process and should be handled with care.
A-4 Electrophoresis pollution
---- Replace the electrophoresis buffer and make sure the consumables and Loading Buffer are free of RNase contamination.
A-5 Too much loading for electrophoresis
---- Reduce the amount of sample loading, the loading of each well should not exceed 2 μg.
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- Q:
DNA contamination
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A-1 Sample amount is too large
---- Reduce sample amount.
A-2 Some samples have high DNA content and can be treated with DNase.
----Perform RNase-Free DNase treatment to the obtained RNA solution, and the RNA can be directly used for subsequent experiments after treatment, or can be further purified by RNA purification kits.
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- Q:
How to remove RNase from experimental consumables and glasswares?
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For glasswares, baked at 150°C for 4 h. For plastic containers, immersed in 0.5 M NaOH for 10 min, then thoroughly rinsed with RNase-free water and then sterilize to completely remove RNase. The reagents or solutions used in the experiment, especially water, must be free of RNase. Use RNase-free water for all reagent preparations (add water to a clean glass bottle, add DEPC to a final concentration of 0.1% (V/V), shake overnight and autoclave).
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